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37 The EFSA Journal (2005) 182, 1-22 Opinion of the Scientific Panel on Genetically Modified Organisms on an application (reference EFSA-GMO-NL ) for the placing on the market of insect-tolerant genetically modified maize 1507, for food use, under Regulation (EC) No 1829/2003 from Pioneer Hi-Bred International/Mycogen Seeds 1 (Question No EFSA-Q ) Opinion adopted on 19 January 2005 SUMMARY This document provides an opinion of the Scientific Panel on Genetically Modified Organisms (GMO Panel) of the European Food Safety Authority (EFSA) on 1507 maize, genetically modified to provide protection against specific lepidopteran pests. The maize also contains a gene providing tolerance to the herbicide glufosinate. In delivering its opinion the Panel considered the application, additional information provided by the applicant and comments submitted by the Member States. Further information from other applications for placing 1507 maize on the market under current regulatory procedures were taken into account where appropriate, as were comments from the Member States. The information from other applications were notification C/ES/01/01 for cultivation, import, processing and use as any other maize (excluding food uses) and notification C/NL/00/10 for import and processing. For regulatory reasons the latter applications resulted in separate opinions maize was assessed with reference to its intended use employing the appropriate principles as described in the Guidance Document of the Scientific Panel on Genetically Modified Organisms for the Risk Assessment of Genetically Modified Plants and Derived Food and Feed. The scientific assessment included examination of the DNA inserted into 1507 maize and the nature and safety of the target proteins produced by the transgenic plants with respect to toxicology and allergenicity. Furthermore, a comparative analysis of agronomic traits and composition was undertaken and the safety of the whole food was evaluated. A nutritional and an environmental assessment, including monitoring plan, were both undertaken maize has been developed for protection against specific lepidopteran pests such as the European corn borer (Ostrinia nubilalis) and Sesamia spp. and for tolerance to the herbicide glufosinate. Insect resistance is achieved by production of a truncated Cry1F protein from Bacillus thuringiensis ssp. aizawai and tolerance to the herbicide is conferred by a phosphinothricin-n-acetyltransferase (PAT) from Streptomyces viridochromogenes. Maize embryos were transformed by particle bombardment to transfer a DNA fragment containing these two genes. As a result of the genetic modification, the 1507 event contains an insert bearing both cry1f and pat genes, under the control of the maize ubiquitin and the 35S promoters, respectively. 1 For citation purposes: Opinion of the Scientific Panel on Genetically Modified Organisms on an application (reference EFSA-GMO- NL ) for the placing on the market of insect-tolerant genetically modified maize 1507, for food use, under Regulation (EC) No 1829/2003 from Pioneer Hi-Bred International/Mycogen Seeds, The EFSA Journal (2005) 182,

38 The EFSA Journal (2005) 182, 1-22 Molecular analysis showed that 1507 maize contains one copy of the DNA fragment used for transformation and that this is present at a single locus in the nuclear genome of the GM plant. The complete DNA sequence of the insert was provided. In addition to the intact genes, the insert in 1507 maize includes DNA sequences originating from the fragment used for transformation as well as maize chloroplast and nuclear genome sequences at both ends of the inserted sequence. While these sequences may have resulted from the transformation process (insertional events), there were no indications that these additional fragments would result in the transcription of new RNA other than the mrnas transcribed from the cry1f and pat genes. In the unlikely event that this does occur, bioinformatics analysis showed that any resulting peptides or proteins would have no homology to known toxins or allergens. Analysis of DNA sequences flanking both ends of the insert shows that they correspond to maize genomic DNA. Analysis of kernel chemical composition from field trials in South America and Europe showed that 1507 maize was substantially equivalent to its non-gm comparator. Furthermore, appropriate animal feeding trials indicated that 1507 maize is nutritionally equivalent to its non- GM comparator. Application EFSA-GMO-NL only concerns food uses for 1507 maize. Therefore, there is no requirement for scientific information on possible environmental effects associated with the cultivation of the GM maize. The GMO Panel agrees that unintended environmental effects due to the establishment and spread of GM maize will not be different from those of maize bred traditionally. The monitoring plan provided by the applicant is in line with the intended uses for the GMO. In conclusion, the GMO Panel considers that the information available for 1507 maize addresses the outstanding questions raised by the Member States and considers that 1507 maize will not have an adverse effect on human and animal health or the environment in the context of its proposed use. This scientific opinion corresponds to the risk assessment report requested under Article 6(6) of Regulation (EC) No 1829/2003 and will be part of the overall opinion as required by Regulation (EC) No 1829/2003. Key words: GMO, maize, Zea mays, 1507, insect protection, Cry1F, PAT, food safety, human health, cultivation, environment, import, Regulation (EC) 258/97, Regulation (EC) 1829/2003, Directive 90/220/EEC, Directive 2001/18/EC. 2

39 The EFSA Journal (2005) 182, 1-22 TABLE OF CONTENTS SUMMARY... 1 BACKGROUND... 3 TERMS OF REFERENCE... 4 ASSESSMENT... 4 CONCLUSIONS AND RECOMMENDATIONS DOCUMENTATION PROVIDED TO EFSA REFERENCES SCIENTIFIC PANEL MEMBERS ACKNOWLEDGEMENT BACKGROUND On 10 June 2004 EFSA received from the Dutch Competent Authority an application submitted by Pioneer Overseas Corporation and Mycogen Seeds within the framework of Regulation (EC) No 1829/2003 on genetically modified food and feed (EC, 2003). The application was originally submitted to the Dutch Competent Authority under Article 4 of Regulation (EC) No 258/97 concerning novel foods and novel foods ingredients (EC, 1997) and covers foods consisting of or derived from genetically modified maize As indicated by the Dutch Competent Authority, the initial assessment was not finalised by 18 April In accordance with Article 46(1) of Regulation (EC) No 1829/2003, the application had, therefore, to be transformed into an application under Regulation (EC) No 1829/2003, following the procedures laid down in Regulation (EC) No 641/2004 (EC, 2004a). According to Article 5(5) of the latter Regulation, the transformed application shall be further processed as any other application under Article 5 of Regulation (EC) No 1829/2003. The application was named EFSA-GMO-NL Member States and the Commission were informed about the transformed application and the summary of the dossier was made publicly available on the EFSA website 2. EFSA initiated a formal review of the application immediately, to check compliance of the dossier submitted with the requirements laid down in Article 5(3) of Regulation (EC) No 1829/2003. After receipt of additional information from the applicant on 20 August 2004 (requested by letter dated 9 August 2004), EFSA declared the application as valid and started the clock in accordance with Article 6(1) Regulation (EC) No 1829/2003 on 3 September As initial steps in the administrative procedures and risk assessment, EFSA made the valid application available to the Member States and the Commission and consulted nominated risk assessment bodies of the Member States, including the national Competent Authorities within the meaning of Directive 2001/18/EC following the requirements of Article 6(4) Regulation (EC) No 1829/2003, to request their comments on the safety assessment of the genetically modified food. The Member State bodies had three months after the date of receipt of the request (until 3 December 2004) within which to make their opinion known. All comments were evaluated by the GMO Panel and taken into consideration for the further risk assessment. Comments on risk management issues, such as co-existence of different agronomic systems, were excluded from further considerations. In delivering its opinion, the Panel considered the application, additional information provided by the applicant and comments submitted by the Member States. Further information from other applications for placing 1507 maize on the market under current regulatory procedures were taken into account where appropriate, as were comments from the Member States. The

40 The EFSA Journal (2005) 182, 1-22 information from other applications referred to notification C/ES/01/01 for cultivation, import, processing and use as any other maize (excluding food uses) and notification C/NL/00/10 for import and processing. For regulatory reasons the latter applications resulted in separate opinions (EFSA, 2004b; EFSA, 2005). In accordance with Article 6(1) of Regulation (EC) No 1829/2003 EFSA shall, in giving its opinion to the Commission, the Member States and the applicant, endeavour to respect a time limit of six months as from the receipt of a valid application. Apart from the requirements listed in Article 6(5) of Regulation (EC) No 1829/2003, the EFSA opinion shall include a report describing the assessment of the food and stating the reasons for its opinion and the information on which its opinion is based. This document is to be seen as the report requested under Article 6(6) of Regulation (EC) No 1829/2003 and thus will be part of the overall opinion as required by Regulation (EC) No 1829/2003. TERMS OF REFERENCE The GMO Panel was requested, in accordance with Article 6(6) of Regulation (EC) No 1829/2003, to carry out a scientific assessment of the genetically modified maize 1507 for food use. Where applicable, any conditions or restrictions which should be imposed on the placing on the market and/or specific conditions or restrictions for use and handling, including post-market monitoring requirements based on the outcome of the risk assessment and, in the case of GMOs or food containing or consisting of GMOs, conditions for the protection of particular ecosystems/environment and/or geographical areas should be indicated in accordance with Article 6(5)(e) of Regulation (EC) No 1829/2003. The Panel was not requested to give an opinion on information required under Annex II to the Cartagena Protocol. The Panel did also not consider proposals for labelling and methods of detection which are matters related to risk management. The latter would include information on sampling and the identification of the specific transformation event in the food and/or foods produced from it. ASSESSMENT 1. Introduction GM 1507 maize was assessed with reference to its intended use and the appropriate principles described in the Guidance Document of the Scientific Panel on Genetically Modified Organisms for the Risk Assessment of Genetically Modified Plants and Derived Food and Feed (EFSA, 2004c). In its evaluation the Panel also considered the issues that were raised by Member States during the initial assessment of the applications introduced under Directive 2001/18/EC and Regulation (EC) 258/97. The assessment presented here is based on the information provided in all available applications relating to GM 1507 maize submitted in the EU including additional information from the applicant in reply to Member States questions. 4

41 The EFSA Journal (2005) 182, Molecular characterisation 2.1. Issues raised by Member States (1) PCR analysis was requested to demonstrate the continuity of the DNA on both sides of the insert in comparison to the recipient plant; (2) a question over the presence of the detected sequences on both sides of the insert giving rise to instabilities of the insert was raised; (3) a question over the existence of a secondary insertion site detectable by Southern analysis was raised; (4) the possibility that very high levels of Cry1F toxin accumulated in specific tissues not subjected to analysis and which might be missed in the analyses was presented Evaluation of relevant scientific data Transformation process and vector constructs Embryogenic cells of Pioneer Hi-II maize were transformed using particle acceleration technology with tungsten particles coated with a purified linear fragment PHI8999A derived from plasmid PHP8999. For this purpose two restriction fragments of 6235 bp and 3269 bp were produced through Pme I-digestion of PHP8999. The larger fragment, named PHI8999A, was purified after agarose gel electrophoresis and the smaller fragment was discarded. DNA fragment PHI8999A contains two adjacent plant gene expression cassettes. The first contains a truncated cry1f gene derived from the Bacillus thuringiensis ssp. aizawai sequence (Chambers et al., 1991). The coding sequence is regulated by a maize ubiquitin promoter and a maize ubiquitin intron sequence introduced upstream of the cry1f sequence. The 3 terminator sequence used is from the Agrobacterium tumefaciens mannopine synthase gene. The second expression cassette contains the pat gene from Streptomyces viridochromogenes (OECD, 1999) which is regulated by a CaMV 35S promoter and terminator. The coding sequence of both genes has been optimised to achieve a high level of expression in maize Transgenic constructs in the genetically modified plant Southern transfer and hybridisation analysis showed the presence of a single insertion locus (comprising a complex structure of different fragments). The absence of vector backbone in the 1507 plants has been confirmed by Southern blotting using probes that cover the entire discarded 3269 bp fragment. The nucleotide sequence of the insert in maize event 1507 has been determined in its entirety, as have sequences of the plant genome adjacent to the 3 and 5 sequences of the insert. Sequence analysis indicates that the insert comprises one almost complete copy 3 of fragment PHI8999A without internal rearrangements. Both cry1f and pat gene cassettes are intact within the transgenic event and the DNA sequences of the genes are identical to those in the original plasmid. The proteins produced in the GM plants are the ones intended, including a leucine residue (replacing a phenylalanine) at position 604 (of 605 amino acids in total) of Cry1F. This modification was introduced to create a specific restriction site for cloning purposes. Southern analysis using a cry1f probe revealed the presence of two cry1f inserts. The first represents the intact gene from the expression cassette. The second insert is a truncated cry1f fragment of 335bp, which is located at the 5 end of the insertion locus. In addition, analysis of 3 Base pairs 1-10 at the 5 end and base pairs at the 3 end are missing. Both missing parts represent polylinker regions of fragment PHI8999A. 5

42 The EFSA Journal (2005) 182, 1-22 the sequences adjacent to the insert of fragment PHI8999A revealed DNA fragments that correspond to small segments from PHI8999A, including incomplete sequences from the pat gene, the maize ubiquitin promoter and the mannopine synthase terminator from Agrobacterium. Furthermore, different fragments of chloroplast DNA and a number of sequences with similarity to retrotransposons are also present in the border region of the insert. PCR analyses indicated that the fragments in the flanking regions can also be found in the recipient line (Hi-II). No data documenting the intactness of the insertion site were shown. Therefore, a direct comparison of the insertion locus and the respective site in the recipient plant is not possible. Sequences found in the border regions showed a high degree of similarity to retrotransposon-like sequences that are considered to be very abundant throughout the maize genome. The design of PCR primers to provide unequivocal evidence that sequences detected in the flanking regions of the 1507 insert are also to be found as continuous sequences in the recipient plant is in general technically difficult. Thus, it cannot be assumed that DNA deletions have not occurred during the transformation process. There is, however, no indication that such a deletion produces any phenotypic effect in the transformed maize line (see Section 3.) Information on the expression of the insert Expression analysis of the Cry1F and the PAT proteins were carried out by Western analysis and ELISA. The tissues and plant samples examined were leaf, pollen, silk, stalk, whole plant and grain. The Cry1F protein was found in all tissues examined while the PAT protein could be detected only in leaf and whole plant. Cry1F Western analyses with protein extracts from different plant tissues revealed a double band (65 to 68 kda) in the range of the predicted size of 66 kda which corresponds to the microbially produced Cry1F protein control. The smaller band detected in the 1507 protein extract is assumed to be the result of a limited N-terminal processing of the full size 1507 Cry1F protein during the extraction process by a plant protease with trypsin-like specificity. This assumption is supported by results from N-terminal amino acid sequencing of the protein which revealed a putative trypsin cleavage site starting at amino acid 28 (of 605) of the Cry1F protein. As no further bands were detected by Western analysis there is no evidence that unintended Cry1F-fusion proteins are expressed in 1507 maize. As additional information, the applicant submitted tables including recalculated the data from Cry1F ELISA experiments. The data are presented on a ng Cry1F protein/mg tissue dry weight basis and show that the expression values fall within the same order of magnitude for cultivation in different years and at different locations. Maximum expression (on a tissue dry weight basis) was found in pollen (average 20.0 and maximum 29.3 ng Cry1F protein/mg tissue dry weight). The values for whole plant extracts ranged between 1.0 and 6.9 ng Cry1F protein/mg tissue dry weight and for kernels between 1.2 to 3.1 ng Cry1F protein/mg tissue dry weight. The expression of Cry1F was not influenced by the application of glufosinate. Measurable expression levels of PAT protein were only found in leaves (<LOD pg/µg TEP 5 ) and whole plant extracts (<LOD 38.0 pg/µg TEP) where the mean value for leaf was 42.0 pg/µg TEP and that for whole plant was below LOD. For kernels, all results were below LOD. Western analysis of PAT protein in leaves revealed only two bands of the expected size (ca. 22 kda and 43 kda [putative homodimer]). This indicates that no partial PAT proteins or fusion proteins were present at detectable levels. 4 LOD = limit of detection 5 TEP = total extractable protein 6

43 The EFSA Journal (2005) 182, 1-22 Bioinformatics analysis of the insert sequence indicates the presence, in addition to the two intended transcripts detected in the transgenic plant, of one further ORF of more than 300 bp length (ORF4: 630 bp) on fragment PHI8999A and a number of other ORFs (including ORF3, which is 753 bp long) spanning the junctions between maize DNA and DNA originating from the transformation fragment. This raises the possibility that new putative fusion proteins could be produced. A detailed analysis of the potential gene expression is provided for the two sequences longer than 300 bp (ORF3 and ORF4). No transcript corresponding to ORF3 was detected either by Northern or by RT-PCR analysis in experiments with mrna from developing kernels. Northern analysis revealed no expression of ORF4 but a weak signal was detected using RT-PCR, which also indicated that the detected mrna originates from a read-through product of the cry1f gene. In the very unlikely event that a protein were expressed from ORF4 on the read-through mrna by using an alternative translation start codon or indeed if any of the other ORFs were transcribed and translated at a very low level, no adverse effects are expected as bioinformatics analysis revealed no significant homologies with known allergens. No known allergenic, toxic or gluten sensitive enteropathy-related proteins are encoded by these ORFs Inheritance and stability of inserted DNA Event 1507 was produced in maize line Hi-II. The event was transferred to a Pioneer elite inbred line and the resulting plants backcrossed to the elite line for six generations. The Mendelian inheritance pattern of the traits was assessed together with the physical linkage of the target genes in resulting progeny. Southern blots and maintenance of the phenotype indicated genetic and phenotypic stability of the transgenic line and their progeny over several generations. No instability of the DNA sequences flanking the insert was observed Conclusion GM maize line 1507 was generated through particle bombardment transformation of maize line Hi-II. Detailed molecular analysis of the insert and Mendelian inheritance of the trait indicated that one copy of fragment PHI8999A used for the transformation was inserted stably over several generations at a single locus in the maize nuclear genome. The inserted fragment is flanked by several fragments originating from the recipient maize plant chloroplast and nuclear genome and from fragment PHI8999A. Evidence that the maize genomic DNA was contiguous with the flanking regions of the insert was not provided. The possibility of undetected deletions at the insertion site caused by the transformation process has been considered. The Panel is of the opinion that it is very unlikely that putative deletions or rearrangements at the insertion locus would result in undiscovered adverse effects. Firstly, a large proportion of the maize genome consists of non-coding sequences. Secondly, other elements of the overall risk assessment (see data provided in Section 3) show no indication of any unintended adverse effects. Thirdly deleted components will, in most cases, be complemented in commercial hybrids. In conclusion, the Panel is of the opinion that the transgenic insert in 1507 maize was analysed and described sufficiently. None of the DNA stretches including the chloroplast DNA sequences detected in the insert region provide grounds for specific concern. The intended expression of the PAT and Cry1F proteins was demonstrated and the expression levels were shown to be in the same range for different locations and growing seasons. The detection of a read-through mrna comprising ORF4 sequences was shown. Bioinformatics assessment provided no indication that the development of allergenic or toxic products would arise in the very unlikely event that the read-through mrna is translated to the respective protein. 7

44 The EFSA Journal (2005) 182, 1-22 Stability of inheritance of the newly inserted DNA and of the expression of the genes that code for Cry1F and PAT proteins in the transgenic plants was demonstrated. 3. Comparative analysis 3.1. Issues raised by Member States (1) Additional data on lignin content were requested, based upon literature data indicating that these levels would be increased in transgenic maize lines expressing B. thuringiensis insecticidal proteins; (2) it was questioned whether levels of Cry1F in tissues of 1507 maize were significantly different over the locations and years; (3) data were requested on the levels of additional chemical substances including supplementary heavy metals, vitamins, and secondary metabolites Evaluation of relevant scientific data Choice of comparator and production of material for the compositional assessment 1507 maize was compared with control hybrids that had not been genetically modified and that had background genetics representative of 1507 maize, except for the inserted genes. Whole crops and maize tissues, including ears with kernels, were collected for compositional analysis from field trials. These field trials occurred during three seasons and at different locations (six locations in Chile ( ), three locations in France and Italy (1999), and six locations in France, Italy and Bulgaria (2000). Maize plants in Chilean field trials were all treated with glufosinate, while those in the European field trials were split into treated and untreated groups Compositional analysis For each season, the results of compositional analyses were provided for the individual and the combined locations. The proximate and mineral analyses (fat, protein, acid detergent fibre (ADF), neutral detergent fibre (NDF), ash, carbohydrate, phosphorus, and calcium) of forage from maize line 1507 (glufosinate-treated and untreated) were comparable to forage from the non-transformed version of the hybrid and within typical ranges reported in literature for commercial maize hybrids. Statistically significant differences were occasionally observed in some GM plants, for example increased overall levels of carbohydrates and decreased levels of fat in forage of maize line 1507 (both sprayed and non-sprayed) in the 2000 season. However, there were no differences that were consistently observed over years and at each location. The compositional analysis of kernels of 1507 maize hybrid and its control included proximate analyses (as for forage above), fatty acid composition [palmitic acid (16:0), stearic acid (18:0), oleic acid (18:1), linoleic acid (18:2), and linolenic acid (18:3)], amino acids (twelve essential and six non-essential amino acids), minerals (calcium, copper, iron, magnesium, manganese, phosphorus, potassium, sodium, and zinc), vitamins (vitamin B1, vitamin B2, folic acid, and total tocopherols), secondary metabolites (inositol, raffinose, furfural, p-coumaric acid, and ferulic acid), and anti-nutrients (phytic acid and trypsin inhibitor). Kernels from the 2000 season were analysed additionally for crude fibre, arachidic acid, provitamin A, and vitamin E. 8

45 The EFSA Journal (2005) 182, 1-22 In summary, the analysis of nutrient composition of kernels from maize line 1507 (glufosinatetreated and non-treated) occasionally revealed statistically significant differences in some compounds. For example, kernels of 1507 maize contained higher overall levels of potassium, linoleic acid, linolenic acid, and tocopherols, as well as lower levels of fat, manganese, stearic acid, oleic acid, cysteine, methionine, and vitamin B1, than control kernels in the season. The levels of protein, amino acids (Ala, Asp, Glu, Gly, His, Leu, Phe, Pro, Ser, Thr, Tyr, and Val), and potassium were increased, while the level of vitamin B2 was decreased, in kernels of 1507 maize (both sprayed and non-sprayed) compared with control kernels in In the 2000 season, ash, amino acids (Ala, Phe, Tyr), and potassium were increased, while manganese was decreased in kernels of maize line 1507 (both sprayed and non-sprayed) compared with controls. Across locations and between years, however, there were no consistent statistically significant differences. All analytical data were either very close to or within the ranges published in the literature. It has been suggested that lignin levels might be increased in transgenic maize lines expressing B. thuringiensis insecticidal proteins (Saxena & Stotzky, 2001; Flores et al., 2005). However, a broader and more extensive study on lignin content in Bt-maize does not support this conclusion (Jung & Sheaffer, 2004). In addition, as mentioned above, the levels of ADF and NDF, which comprise lignin, in forage of 1507 maize were comparable with those in control maize and within the background range. Moreover, similar levels were observed for the lignin precursors p- coumaric acid and ferulic acid in kernels of 1507 maize and control maize, except for a small but statistically significant difference in p-coumaric acid between sprayed 1507 maize and control maize in the 2000 season. Aside from minor modifications, the selection of compounds analysed followed the recommendations of OECD (OECD, 2002). During the Member State consultation under Article 6.4 of Regulation (EC) No. 1829/2003, it was suggested that additional compounds, including certain heavy metals, vitamins, and secondary metabolites, should be analysed. The Panel is of the opinion, however, that such additional information would not add value to the data that had already been provided, given, among other things, the high variability of the levels of some compounds like selenium and DIMBOA 6, due to environmental conditions or the stage of plant development Agronomic traits and GM phenotype Studies of plant biology and canopy morphology complemented extensive agronomic data and confirmed the similarity of 1507 maize to its non-transgenic counterpart. During field trials over several seasons and at different locations (USA in 1999, France, Italy, and Bulgaria in 2000, Spain in 2002) extensive agronomic data (germination as early stand counts, visual ratings of development, accumulated heat units to pollen shed and silking, stalk and root lodging, plant height, ear height, final population, date/time of leaf senescence, disease incidence, insect damage, grain moisture and density) were collected and confirmed the similarity of 1507 maize phenotype to its non-transgenic counterpart. Slight differences in accumulated heat units to pollen shed and silking under infestation were reported and are regarded as indicative of small differences in the genetic background of the GM- and non-gm-hybrids. No differences in the general appearance of the plants or other phenotypical differences that would indicate unexpected pleiotropic effects of the genetic modification were found. 6 DIMBOA = 2,4-dihydroxy-7-methoxy-1,4-benzoxazin-3-one, a metabolite naturally formed by maize plants. 9

46 The EFSA Journal (2005) 182, Conclusion Based on the results of compositional analysis of samples from a representative range of environments and grown in three seasons, it is concluded that forage and kernels of 1507 maize are compositionally equivalent to those of conventional maize, except for the presence of Cry1F and PAT proteins in 1507 maize. In addition, experimental field trials in the USA and Europe did not show indications for unexpected changes of agronomic characteristics and performance. 4. Food/feed safety assessment 4.1. Issues raised by Member States (1) Bioinformatic analysis was requested to compare the conformations of MR872 (microbially produced, trypsinised Bt toxin) and the plant-expressed Cry1F protein; (2) it was argued that the Cry1F produced by plants might differ from the Bt toxin produced by bacteria, e.g. with regard to posttranslational modifications besides glycosylation; (3) further animal feeding studies, including tests on ruminants, laying hens, pigs, fish, and crustaceans, with whole products, including forage, derived from 1507 maize were requested; (4) additional toxicological studies comprising various trials, including chronic testing were requested; (5) clarification on the decreased average eosinophil counts in female rats fed diets containing 33% 1507 maize was requested Evaluation of relevant scientific data Product description and intended use Application EFSA-GMO-NL covers foods consisting of or derived from genetically modified maize The feed uses of 1507 maize and the aspects of cultivation, import and processing are covered by other applications 7. Maize plants and kernels are used mainly for animal feed but, on a smaller scale, sweet maize kernels are also used for direct human consumption. Products from maize kernels such as flour, starch (also transformed to sweeteners e.g. as syrups) and maize germ oil can be regarded as important base materials for food production. As the modification in 1507 maize is only intended to improve the agronomic performance but not to influence nutritional aspects, production processes and overall use of maize as a crop are not expected to be influenced as a result of the introduction of the GM plants to the market Stability during processing Experimental fish feed containing 38.7% maize meal was prepared in order to test the stability of Cry1F during processing. The Cry1F level in transgenic maize kernels was ng/mg tissue dry weight prior to processing. The production of fish feed included an extrusion step, exposing feed ingredients to high pressure and temperature. Cry1F was not detectable in the final product, as established through an insect bioassay and immuno-assay (ELISA LOD = 0.04 ng/mg tissue dry weight). 7 Notifications C/ES/01/01 and C/NL/00/10 submitted under Directive 2001/18/EC 10

47 The EFSA Journal (2005) 182, 1-22 In addition, the thermostability of recombinant Cry1F protein produced by Pseudomonas fluorescens at elevated temperatures was assessed by heating solutions of 1.3 ppm Cry1F in phosphate buffer ph 7.5 at 60, 75, or 90ºC for 30 minutes. Samples were taken from these solutions and added to feed used in a bioassay for insecticidal activity on tobacco budworm (Heliothis virescens). It was thus observed that the Cry1F proteins heated at 75 and 90ºC had lost their insecticidal activity Toxicology Cry1F and PAT proteins used for safety assessment Given the low expression levels of Cry1F in 1507 maize, the applicant decided to use a trypsinised microbial analogue, MR872, of the truncated Cry1F protein expressed in maize line 1507 for safety testing. To this end, a fusion protein consisting of the non-truncated Cry1F (N-terminal) linked to Cry1Ab (C-terminal) was produced by recombinant Pseudomonas fluorescens. Trypsin cleavage sites in Cry1F are located between residues 28-29, 31-32, and Enzymatic cleavage with trypsin of the fusion protein yielded a core protein, MR872, identical to the truncated Cry1F protein expressed in 1507 maize, except for i) phenylalanine (Phe) instead of leucine (Leu) at position 604 and ii) a C-terminal extension of trypsinised MR872 with seven amino acid residues ( , Ala-Glu-Tyr-Asp-Leu-Glu-Arg). With regard to the conformation of Cry1F, it is considered unlikely that the substitution at position 604 would lead to conformational changes because both Phe and Leu are amino acids with hydrophobic side chains. The extension of the trypsinised MR872 protein with seven amino acids at the C-terminus of domain III is also present in native Cry1F from B. thuringiensis, as well as in other Cry proteins. Comparison of the crystal structure of Cry1Aa containing this extension (Grochulski et al., 1995) with that of Cry3A lacking this extension (Li et al., 1991) does not indicate differences in the overall structure of Domain III. It is therefore unlikely that this extension would affect the functional, toxicological, or allergenic properties of the protein. Both bacterially produced Cry1F and plant-expressed Cry1F isolated from leaves and kernels of 1507 maize displayed a prominent 65 kda band on Western blots, which corresponds to the N-terminally processed form of plant-expressed Cry1F as mentioned in Section Glycosylation was analysed after SDS PAGE using a commercial staining kit. The results demonstrate that the plant-expressed Cry1F is not glycosylated. Moreover, MALDI-TOF mass spectrometry was performed on trypsin-digests of the recombinant Cry1F proteins produced by transgenic P. fluorescens and 1507 maize and separated by electrophoresis. Fragments were observed in the spectra of both types of Cry1F protein that concurred with the predicted masses of peptides derived from trypsin digestion, covering percent of the total protein sequence (605 amino acids) encoded by the cry1f transgene in 1507 maize in various experiments. Data provided by the applicant on insect bioassays with recombinant Cry1F show no notable differences between preparations of this protein isolated from transgenic maize event 1360 (modified with Cry1F) and P. fluorescens. Taking into account all the evidence provided, the Panel is of the opinion that the trypsinised MR872 analogue is an appropriate substitute of the Cry1F protein expressed in 1507 maize for safety testing. Bacterially produced recombinant PAT showed the same electrophoretic mobility as PAT expressed in 1507 maize during Western blotting. As noted above, levels of PAT were not quantifiable in kernels of 1507 maize. 11

48 The EFSA Journal (2005) 182, Toxicological assessment of expressed novel proteins in 1507 maize (a) Acute oral toxicity An acute oral study was performed in albino mice dosed with 576 mg truncated Cry1F/kg bodyweight (5050 mg/kg test material containing 11.4% Cry1F). No effects related to the administration of Cry1F were noted on bodyweight, gross necropsy, and mortality 14 days after the administration, except for one incidental finding out of 10 of lack of body weight gain between days 7 and 14. For PAT, a study was performed, in which mice received 5000 mg PAT/kg bodyweight (equal to 6000 mg test material/kg). After two weeks, no effects on bodyweight and gross pathology were noted. (b) Degradation in simulated digestive fluids The trypsin-resistant core of the microbially produced Cry1F protein was rapidly degraded (<1 minute) in simulated gastric fluid at a Cry1F/pepsin molar ratio of 188:1 and 1:22. In the SDS PAGE gels of the incubation mixture, a 10-kDa band was visible that was relatively stable during the period of the experiments. This was probably a contamination of the microbial Cry1F preparation, as it was not detected in Western analysis with anti-cry1f immune sera. In simulated intestinal fluid (pancreatin), the trypsin-resistant Cry1F core protein proved stable over the entire exposure of 120 minutes. For degradation of the PAT protein, reference is made to previous studies in which PAT was degraded within 5 seconds in simulated gastric fluid Toxicological assessment of new constituents other than proteins Since no new constituents other than the above mentioned proteins were expressed in 1507 maize, nor were levels of endogenous compounds altered, a toxicological assessment is not applicable Toxicological assessment of the whole GM food/feed Subchronic oral toxicity A 90-day oral toxicity study has been performed on rats in five groups (12 animals/sex/group) fed diets containing 1507 maize (11 and 33%), a non transgenic control line with comparable genetic background (11 and 33%), and another non transgenic maize line as reference (33%). The diets were analysed for nutrients, antinutrients, mycotoxins, pesticides, heavy metals, transgenic DNA, and Cry1F (insect bioassay). Kernels used in this study were obtained from 1507 maize plants that had not been treated with glufosinate. The measurements on animals included feed consumption, body weight, clinical pathology (serum, blood, urine), and anatomical pathology (organ weights, histopathology). A statistically significant increase in feed consumption was observed in male rats fed 33% 1507 maize compared with rats fed control maize, but not to those fed the reference maize (27.5 ± 2.6, 25.7 ± 1.7, and 27.3 ± 1.7 g per day, respectively). This effect is therefore not considered to pose concerns over the safety and nutritional value of 1507 maize. In addition, serum counts of eosinophil leukocytes were statistically significantly decreased in female rats fed 33% 1507 maize compared with those fed 33% near isogenic control and reference maize. 12

49 The EFSA Journal (2005) 182, 1-22 The observed differences were not considered to be biologically relevant, since (1) it was observed in one sex only, (2) this was an isolated finding in a series of haematological parameters, and (3) the inherent variability of the measured parameter. A number of histopathological changes were observed, in particular inflammation of the liver, nephropathy, and cardiomyopathy (kidney and heart damage) in animals of both sexes. To a lesser degree, inflammation of the prostate in males and the pancreas in females, fatty change in the liver of females, and atrophy of the pancreas in males were observed. These effects were not linked to the test-substance, since their incidences were not elevated substantially in the animals fed 1507 maize compared to control animals. This study, on the basis of presented results, is considered satisfactory and does not raise concerns over the safety of 1507 maize Allergenicity The strategies in assessing the allergenic risk concentrate on characterisation of the source of the recombinant protein, the potential of the newly expressed protein to induce sensitisation or to elicit allergic reactions in already sensitised persons and whether the transformation may have altered the allergenic properties of the modified food. A weight of evidence approach is recommended, taking into account all of the information obtained with various test methods, since no single experimental method yields decisive evidence for allergenicity (EFSA, 2004c; CAC, 2003) Assessment of allergenicity of the newly expressed proteins The PAT protein has been previously evaluated for its safety in the frame of other applications for the placing of PAT-expressing GM crops on the market. The potential allergenicity of the transgenic Cry1F protein and of the theoretical expression products of ORF4 (within the PHI8999A copy of the insert), and 24 ORFs (including ORF3) coding for putative fusion proteins in the regions adjacent to the PHI8999A copy of the insert were considered in this dossier. The amino acid sequence of the Cry1F protein has been compared with the sequences of allergenic proteins compiled in an allergen database 8. This comparison focused on two types of identity between Cry1F and allergens: (1) short linear stretches; with a relevant minimum size of eight contiguous amino acids and (2) overall identity of 80-amino-acid peptides of Cry1F (min. 35% identity relevant). For both types of comparison, the FastA algorithm was applied, with appropriate settings. No outcomes were equal to or exceeded the minimum relevant size. The length of the longest identical short linear stretch, for example, was six amino acids. In addition, comparison of the Cry1F sequence against a general protein database yielded predominantly homologies with other Cry-proteins (e.g. Cry1Ab with 52.4% identity over a 614 residue alignment overlap), except for three proteins from Methanosarcina acetivorans, Saccharomyces cerevisiae, and Sinorhizobium meliloti. These three proteins are not known to be toxic and therefore this result does not indicate any homology of the Cry1F with toxic proteins. Three different linear six-amino acid stretches were found to be shared by Cry1F with allergenic proteins (Der p 7 from house dust mite, beta-1,3-glucanase-like protein from olive, and Can f 3 from dog dander). The EFSA panel is aware of studies that show that using a threshold of six amino acids for identical stretches between a given protein and allergens yields a high number of false positives, i.e. this threshold makes the comparison non-specific. Using a newly 8 Applied update: March comprises 2033 entries compiled from published lists supplemented through a search of public domain protein databases. 13

50 The EFSA Journal (2005) 182, 1-22 developed methodology (Soeria-Atmadja et al., 2004), the Swedish National Food Authority found that for Cry1F, many six-amino acid identities with non-allergenic proteins existed (data not published). Kleter & Peijnenburg (2002) further found that many transgenic proteins shared identical six- and seven-amino acid stretches with allergens. For the identical sequences that Cry1F shared with allergens (the same as found by the applicant) these authors found no indications that they were part of IgE-epitopes. Therefore it is unlikely that these identical stretches within Cry1F would induce allergic reactions. In addition, the highest degree of similarity of 80-residue fragments of Cry1F was 33.8% identity (27 residues) with a pollen allergen (Syr v I) from Syringa vulgaris and with related olive pollen allergens. Because the minimum relevant matches are eight-amino-acid linear sequences and 35% identity of 80-residue fragments, respectively, the search has yielded no outcomes that raise safety concerns for Cry1F. The same methodology to search for short identical and larger similar stretches of homology to the proteins listed in the allergen database has been applied to assess the hypothetical peptides derived from ORF4 (within the copy of the PHI8999A sequence on the insert) and the 24 ORFs (including ORF3) coding for putative fusion proteins in the regions adjacent to the PHI8999A copy on the insert. In addition, the ORF3 and ORF4 sequences were compared with the sequences of a general protein database. For ORF 4, the longest identical short linear stretch, for example, was six amino acids, shared with allergenic proteins from durum wheat (glutenin) and wheat (gamma-gliadin). An 80-residue fragment of ORF4 shared twenty-two identical residues (27.5%) with major hazel pollen allergen Cor a 1. In a comparison of ORF4 to general protein sequences, the protein from ORF VI of Cauliflower Mosaic Virus, followed by proteins from Carnation Etched Ring virus and Plasmodium falciparum, were most identical to the ORF4 sequence. ORF3 shared two identical linear sequences of six amino acids with the allergen Gly m IA from soybean and with the allergens gamma-gliadin and alpha/beta-gliadin from wheat. In addition, an 80-residue fragment of ORF3 shared eighteen identical residues (22.5%) with the allergenic barley alpha amylase/trypsin inhibitor precursor and also with Sin a I allergen from white mustard. The highest scoring identities of the sequence of ORF3 with general protein sequences in a public database were those with chloroplast RNA polymerases of various plants and with phosphinothricin acetyltransferase enzymes. Some of the other 23 ORFs in the flanking regions shared six-amino acid identities with allergens. However, none of these ORFs shared relevant homologies with allergens consisting of identical linear sequences of a minimum of eight-amino acids or 35%-identities of 80-amino acid subsequences. In the comparison of these ORFs with a general protein database, none of the sequences sharing the most relevant identities with the ORFs were known to be toxic. The sequence homologies that have been found, therefore, do not raise concerns over the safety of 1507 maize that would justify additional studies regardless the fact that those ORFs are very unlikely to be transcribed and/or translated into peptides or proteins. The degradation of gene products during processing at high temperature and in simulated digestive fluids, which is also relevant for the assessment of potential allergenicity, has been discussed in Sections and Based on all information made available, the Panel considers that the newly expressed proteins are not likely to be allergenic. 14

51 The EFSA Journal (2005) 182, Assessment of allergenicity of the whole GM plant or crop Allergenicity of the whole crop could be increased as an unintended effect of the random insertion of the transgene in the genome of the recipient, for example through qualitative or quantitative modifications of the pattern of expression of endogenous proteins. This issue does not appear relevant to the Panel since maize is not considered a major allergenic food and possible over-expression of any endogenous protein that is not known to be allergenic would be unlikely to alter the overall allergenicity of the whole plant. The same considerations also apply for exposure by inhalation Nutritional assessment of GM food/feed A 42-day feeding study was carried out with broilers to investigate nutritional equivalency. Diets contained on average 55% dry matter (DM) maize kernels from either the transgenic hybrid 1507 maize, the control hybrid maize Mycogen 7250, and four commercial maize hybrids. Each diet was fed to 35 animals (divided into 7 replicates of 5 animals). No statistically significant differences were observed for mortality, body weight, body weight gain, and feed conversion between the different maize lines. Twenty lactating dairy cows were used in a single cross-over design in which there was 2 x 28- day feeding periods. The aim was to compare the effect of using maize silage and maize kernels derived from transgenic 1507 maize on feed intake and milk production when compared with maize silage and maize kernels derived from non-gm control hybrids. Diets contained on average 43.0% DM maize silage and 22.1% concentrate of which 70.2% was in the form of ground maize. Other feed ingredients included alfalfa hay, soybean meal, and cotton seeds. The diet composition was analysed for proximates, minerals (Ca, P, Mg, K), mycotoxins and silage fermentation products and found to be similar for both treatment groups. Cry1F was detected in transgenic maize kernels and silage. PAT was not detectable in kernels, and ranged from not detectable to slightly above the detection threshold in forage, of 1507 maize. The following measurements were made: (1) Physical (weekly): body weight, condition, temperature, pulse, feed intake; (2) Milk production (daily); (3) Milk composition (weekly): protein, fat, dry matter, lactose, urea N, somatic cell count, Cry1F; (4) Blood analysis (prior to and at the end of both trials): chemical and haematological. One cow was positive for the presence of Cry1F in milk prior to and during both treatments, which can therefore be considered a false positive ELISA-reaction. In conclusion, results showed no significant differences between dietary treatments and indicate nutritional equivalence between the transgenic 1507 maize and the non-gm control Post-market monitoring of GM food/feed 1507 maize is intended to have improved agronomic properties. From a nutritional point of view the maize is equivalent to conventionally bred hybrids. Therefore the GM plants will be used as any other maize and only replace a part of the overall maize products within the European market. The risk assessment concluded that no data have emerged to indicate that maize line 1507 is any less safe than its non-gm comparators. The opinion of the applicant that a postmarket monitoring of the GM food/feed is not necessary is in line with the guidance document of the GMO Panel for the risk assessment of genetically modified plants and derived food and feed (EFSA, 2004c) and is shared by the GMO Panel. 15

52 The EFSA Journal (2005) 182, Conclusion The transgenic Cry1F protein showed no adverse effects in an acute oral mouse study. In addition, Cry1F displayed instability towards conditions that prevailed during the production of fish feed including heating and was rapidly degraded in simulated gastric fluid. The sequence of the transgenic Cry1F did not show any significant similarity with the sequences of known allergens. Neither the hypothetical peptide sequences corresponding to 24 ORFs that are present on the insert in 1507 maize nor ORF4 on fragment PHI8999A show significant similarity to allergens or toxins. With regard to animal studies with the whole product, no oral toxicity of 1507 maize was observed in a 90-day rat study. In addition, nutritional data comprising target animal feeding studies with the whole maize kernel on broilers and dairy cows indicate that 1507 maize is nutritionally equivalent to other conventional maize cultivars. These animal studies therefore further support the findings of the compositional analysis of no effect beyond the intended introduction of the PAT and Cry1F proteins. Based on the data provided, the Panel is of the opinion that there is no need for additional chronic toxicity testing, nor for testing in other target animal species. 5. Environmental risk assessment and monitoring plan 5.1. Issues raised by Member States Concerns were expressed that (1) the environmental risk assessment did not adequately address environmental exposure and (2) that the monitoring plan provided by the applicant was not sufficient Evaluation of relevant scientific data Environmental risk assessment Potential unintended effects on plant fitness due to the genetic modification Application EFSA-GMO-NL is for the placing on the market of foods consisting of or derived from 1507 maize only. Maize is highly domesticated and not generally able to survive in the environment without cultivation. Maize plants are not winter hardy in most parts of Europe, they have lost their ability to release seeds from the cob and they do not occur outside cultivated land in Europe, despite cultivation for many years. In addition, there are no cross-compatible wild relatives in Europe, and gene flow via pollen is largely restricted to neighbouring crops. Maize is a hybrid crop and thus imported grain will be a segregated F2 generation and not as fit as the F1 in case of accidental spillage. Field experiments carried out in France, Italy, Bulgaria and in South America demonstrated that 1507 maize has no altered survival, multiplication or dissemination characteristics. The Panel agrees with the assessment that the likelihood of unintended environmental effects due to the establishment and spread of 1507 maize will be no different to that of traditionally bred maize Potential for gene transfer A prerequisite for any gene transfer is the availability of pathways for the transfer of genetic material, DNA in case of horizontal gene transfer and pollen in case of vertical gene flow through cross-pollination. 16

53 The EFSA Journal (2005) 182, 1-22 Exposure of microorganisms to transgenic DNA derived from GM maize plants takes place in the environment during natural decay of transgenic plant material, such as GM plant parts, in agricultural areas and/or pollen in nearby natural ecosystems as well as in cropped fields. Transgenic DNA is a component of some or most of the food and feed products derived from the GM maize. Therefore microorganisms in the digestive tract of humans and animals (domesticated animals and other animals feeding on fresh and decaying GM plant material) may be exposed to transgenic DNA. Transgenic pollen is shed and distributed from cultivated GM hybrids or from plants resulting from the adventitious presence of GM kernels in conventionally bred maize seeds. A further but less likely pathway of dispersal of transgenic maize pollen is the flowering of volunteer GM maize plants originating from accidental seed spillage during transport and/or processing. For Zea mays any vertical gene transfer is limited to other maize plants as populations of sexually compatible wild relatives of maize are not known in Europe. (a) Plant to bacteria gene transfer Based on present scientific knowledge and elaborated recently in more detail (EFSA, 2004d), gene transfer from GM plants to bacteria under natural conditions is extremely unlikely, and would occur primarily through homologous recombination in microbes. The cry1f gene and the pat gene expressed in the 1507 maize are under the control of eukaryotic promoters with limited if any activity in prokaryotic organisms. Genes under control of prokaryotic regulatory elements conferring the same traits as expressed in the GM plants are widespread in microorganism in natural environments. Taking into account the origin and nature of these genes and the lack of selective pressure in the intestinal tract and/or the environment, the likelihood that horizontal gene transfer would confer selective advantages or increased fitness on microorganisms is very limited. For this reason it is very unlikely that genes from 1507 maize would become established in the genome of microorganism in the environment or human and animal digestive tract. In the very unlikely event that such a horizontal gene transfer would take place, no adverse effects on human and animal health and the environment are expected as no principally new traits would be introduced into microbial communities. (b) Plant to plant gene transfer The extent of cross-pollination to conventionally bred hybrids will mainly depend on the scale of accidental release and/or adventitious presence in conventional seeds. As shown in several field trials there are no indications for an altered ecological fitness of the GM maize in comparison to conventionally bred hybrids with similar genetic background. The herbicide resistance trait can only be regarded as providing a selective advantage where and when glufosinate-ammonium containing herbicides are applied, i.e. mainly on arable land. Insect protection against lepidopteran pests is also not regarded as providing a selective advantage for maize in Europe, as the survivability is mainly limited by the absence of a dormancy phase, susceptibility to fungi and susceptibility to cold climate conditions. Therefore, as for any other maize cultivars, it is considered very unlikely that volunteers could survive until subsequent seasons or would establish undesirable populations under European environmental conditions. 17

54 The EFSA Journal (2005) 182, Potential interactions of the GM plant with non-target organisms There is an issue that gene products, particularly Cry proteins might enter the environment either from the gastrointestinal tracts of animals (manure and faeces), through horizontal gene flow to bacteria or as part of waste waters and/or dusts from food production. Data supplied by the applicant and other literature suggests that most protein would be denatured by enzymatic activity in the gastrointestinal tract so that little Cry toxin would survive to pass out in faeces. There would subsequently be further degradation of proteins in the manure due to microbial processes. Thus amounts of Cry proteins being distributed onto land in manure would be very low, minimising the possibility for exposure of potentially sensitive non-target organisms. The GMO Panel considered possible differences between the plant expressed and the microbially-derived Cry1F protein regarding potential effects on non-target organisms. Equivalence tests showed that the activity and structure of Cry1F proteins derived from plant and microbe are comparable. Furthermore, the amino acid sequence of the biologically active core, immunoreactivity, glycosylation and biological activity were comparable between plantexpressed and microbially-produced protein Monitoring The objectives of a monitoring plan according to Annex VII of Directive 2001/18/EC are to confirm that any assumption regarding the occurrence and impact of potential adverse effects of the GMO, or its use, in the environmental risk assessment are correct and to identify the occurrence of adverse effects of the GMO, or its use, on human health or the environment which were not anticipated in the environmental risk assessment. The scope of the monitoring plan provided by the applicant is in line with the intended uses for the GMO since the environmental risk assessment did not cover cultivation Conclusion Application EFSA-GMO-NL only covers food uses of 1507 maize and thus there is no requirement for scientific information on environmental effects associated with cultivation. Maize is highly domesticated and not able to survive in the environment without cultivation. The Panel agrees that unintended environmental effects due to the adventitious establishment and spread of GM maize will be no different to that of traditionally bred maize. The scope of the monitoring plan provided by the applicant is in line with the intended uses for the GMO since the environmental risk assessment did not cover cultivation. CONCLUSIONS AND RECOMMENDATIONS Maize line 1507 has been developed for protection against lepidopteran pests by expressing the Cry1F Protein and for tolerance to glufosinate by the introduction of a pat gene. The GMO Panel has assessed information provided on molecular inserts within the transgenic event, on the safety of the proteins expressed and on the potential for risks associated with any changes to the nutritional, toxicological and allergenic properties of 1507 maize. Analysis of the chemical composition of the maize and field trial data were also used to assess the potential for changes to safety, nutritional as well as agronomic parameters. No data have emerged to indicate that maize line 1507 is any less safe than its non-gm comparators. 18

55 The EFSA Journal (2005) 182, 1-22 The GMO Panel considers that 1507 maize will have similar impacts as other comparable non- GM maize cultivars on the environment. The only adverse effect identified was the possibility that resistance to Bt toxin might evolve in corn borers exposed to 1507 maize following cultivation for some years. The Panel accepts the monitoring plan developed by the applicant to monitor specifically for resistance in corn borers and recommends that cultivation should be accompanied by appropriate risk management strategies to minimise exposure of both target and non-target insects to Bt toxins. In addition, the Panel accepts in principle the general surveillance plan submitted by the applicant. The GMO Panel is therefore of the opinion that there is no evidence to indicate that placing of maize line 1507 and derived products on the market is likely to cause adverse effects on human or animal health or the environment in the context of its proposed use. The authorisation of the complementary herbicide is not within the remits of this opinion and is covered by other legal frameworks of the EU and Member States. The GMO Panel is of the opinion that, based on the outcome of the risk assessment, no specific conditions or restrictions should be imposed on the placing of 1507 maize on the market for food use. No specific conditions or restrictions for food use and handling, including post-market monitoring requirements regarding the use of 1507 maize for human consumption, are regarded as necessary. Furthermore, there is no need for specific conditions for the protection of particular ecosystems/environment and/or geographical areas. DOCUMENTATION PROVIDED TO EFSA 1. Letter from the Dutch Competent Authority (Ministerie van Volksgezondheid, Welzijn en Sport, Directie Voeding en Gezondheidsbescherming), dated 4 June 2004 concerning the submission to EFSA of application 1507 maize within the framework of Regulation (EC) 1829/2003 (Ref. VGB/VL ). 2. Letter from Guy Van Den Eede (IHCP-JRC-Ispra/Italy), dated 27 July 2004, concerning the completeness check of application EFSA-GMO-NL in accordance with Article 5(3)(i) and (j) and Article 17(3)(i) and (j) of Regulation (EC) 1829/2003 (ref. JRC I06- BGMO/GVDE/D/2004 (56) 19688). 3. Letter from EFSA to applicant, dated 3 September 2004, concerning the Statement of Validity for application EFSA-GMO-NL , 1507 maize submitted under Regulation (EC) 1829/2003 (Ref. SR/KL/jq (2004)631). 4. Submission of the application EFSA-GMO-NL by Pioneer/Mycogen Seeds to EFSA, containing: Part I Part II Part III Part IV Part V Part VI technical dossier summary Cartagena Protocol labelling proposal samples and detection method additional information for GMOs 5. The following application dossiers concerning 1507 maize including assessment reports, the respective Member States comments/objections and additional information submitted by Pioneer/Mycogen Seeds were considered where appropriate: 19

56 The EFSA Journal (2005) 182, 1-22 a. Notification (C/NL/00/10) to market products containing genetically modified organisms in accordance with Directive 2001/18/EC submitted by Pioneer/Mycogen Seeds to EFSA on 26 March b. Application for placing on the market of novel foods and novel food ingredients containing genetically modified organisms in accordance with Regulation (EC) 258/97 submitted by Pioneer/Mycogen Seeds to EFSA on 26 March c. Notification (C/ES/01/01) to market products containing genetically modified organisms in accordance with Directive 2001/18/EC submitted by Pioneer/Mycogen Seeds to EFSA on 19 May REFERENCES CAC, Codex principles and guidelines on foods derived from biotechnology. Joint FAO/WHO Food Standards Programme, Food and Agriculture Organisation, Rome. ftp://ftp.fao.org/codex/standard/en/codextextsbiotechfoods.pdf Chambers, J.A., Jelen, A., Gilbert, M.P., Jany, C.S., Johnson, T.B.& Gawron-Burke, C., Isolation and characterization of a novel insecticidal crystal protein gene from Bacillus thuringiensis sbsp. aizawai. J. Bacteriol., 173(13), EC, Council Directive 91/414/EEC of 15 July 1991 concerning the placing of plant protection products on the market. OJ, L230, EN&numdoc=31991L0414&model=guichett EC, Regulation (EC) No 258/97 of the European Parliament and of the Council of 27 January 1997 concerning novel foods and novel food ingredients OJ, L43, EN&numdoc=31997R0258&model=guichett EC, 2001 Directive 2001/18/EC of the European Parliament and of the Council of 12 March 2001 on the deliberate release into the environment of genetically modified organisms and repealing Council Directive 90/220/EEC. Official Journal of the European Communities, L106, EC, Regulation (EC) No 1829/2003 of the European Parliament and of the Council of 22 September 2003 on genetically modified food and feed. OJ, L268, EFSA, 2004a. Commission Regulation (EC) No 641/2004 of 6 April 2004 on detailed rules for the implementation of Regulation (EC) No 1829/2003 of the European Parliament and of the Council as regards the application for the authorisation of new genetically modified food and feed, the notification of existing products and adventitious or technically unavoidable presence of genetically modified material which has benefited from a favourable risk evaluation. OJ L102,

57 The EFSA Journal (2005) 182, 1-22 EFSA, 2004b. Opinion of the Scientific Panel on Genetically Modified Organisms on a request from the Commission related to the Notification (Reference C/NL/00/10) for the placing on the market of insect-tolerant genetically modified maize 1507, for import and processing, under Part C of Directive 2001/18/EC from Pioneer Hi-Bred International/Mycogen Seeds, The EFSA Journal, 124, EFSA, 2004c. Guidance document of the Scientific Panel on Genetically Modified Organisms for the Risk Assessment of Genetically Modified Plants and Derived Food and Feed. The EFSA Journal 99, EFSA, 2004d. Opinion of the Scientific Panel on Genetically Modified Organisms on the use of antibiotic resistance genes as marker genes in genetically modified plants, The EFSA Journal 48, EFSA, Opinion of the Scientific Panel on Genetically Modified Organisms on a request from the Commission related to the notification (Reference C/ES/01/01) for the placing on the market of insect-tolerant genetically modified maize 1507, for import, feed and industrial processing and cultivation, under Part C of Directive 2001/18/EC from Pioneer Hi-Bred International/Mycogen Seeds, The EFSA Journal (2005) 181, Flores, S., Saxena, D. & Stotzky, G., Transgenic Bt plants decompose less in soil than non- Bt plants. Soil Biology & Biochemistry, in press. Grochulski, P., Masson, L., Borisova, S., Pusztai-Carey, M., Schwartz, J.L., Brousseau, R., & Cygler, M., Bacillus thuringiensis CryIA(a) Insecticidal Toxin: Crystal Structure and Channel Formation. J. Mol. Biol. 254(3), Jung, H.G. & Sheaffer, C.C., Influence of Bt Transgenes on Cell Wall Lignification and Digestibility of Maize Stover for Silage. Crop Science, 44, Kleter, G.A. & Peijnenburg, A.A.C.M., Screening of transgenic proteins expressed in transgenic food crops for the presence of short amino acid sequences identical to potential, IgE-binding linear epitopes of allergens. BMC Structural Biology, 2, 8. ype=pdf Li, J.D., Carroll, J. & Ellar, D.J., Crystal structure of insecticidal delta-endotoxin from Bacillus thuringiensis at 2.5 A resolution. Nature, 353, OECD, Consensus document on general information concerning the genes and their enzymes that confer tolerance to phosphinothricin herbicide. Series on Harmonization of Regulatory Oversight in Biotechnology No. 11. Organisation for Economic Co-operation and Development (OECD), Paris. OECD, Consensus Document on Compositional Considerations for New Varieties of Maize (Zea Mays): Key Food and Feed Nutrients, Anti-nutrients and Secondary Plant Metabolites. Series on the Safety of Novel Foods and Feeds, No. 6, Organisation for Economic Co-operation and Development (OECD), Paris

58 The EFSA Journal (2005) 182, 1-22 Saxena, D. & Stotzky, G., Bt corn has a higher lignin content than non-bt corn. Am. J. Bot., 88, Soeria-Atmadja, D., Zorzet, A., Gustafsson, M &Hammerling, U., Statistical evaluation of local alignment features for prediction of allergenicity using supervised classification algorithms. Int. Arch. Allergy Immunol., 133, SCIENTIFIC PANEL MEMBERS Hans Christer Andersson, Detlef Bartsch, Hans-Joerg Buhk, Howard Davies, Marc De Loose, Michael Gasson, Niels Hendriksen, John Heritage, Sirpa Kärenlampi, Ilona Kryspin-Sørensen, Harry Kuiper, Marco Nuti, Fergal O Gara, Pere Puigdomenech, George Sakellaris, Joachim Schiemann, Willem Seinen, Angela Sessitsch, Jeremy Sweet, Jan Dirk van Elsas and Jean-Michel Wal. ACKNOWLEDGEMENT The GMO Panel wishes to thank Gijs Kleter and Richard Phipps for their contributions to the draft opinion. 22

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60 EUROPEAN COMMISSION DIRECTORATE GENERAL JRC JOINT RESEARCH CENTRE INSTITUTE FOR HEALTH AND CONSUMER PROTECTION COMMUNITY REFERENCE LABORATORY FOR GM FOOD AND FEED Event-specific method for the quantitation of maize line TC1507 using real-time PCR Protocol Method development: Pioneer Hi-Bred International GeneScan Analytics GmbH Method validation: Joint Research Centre European Commission Biotechnology & GMOs Unit

61 Protocol TC1507 Community Reference Laboratory 2/17 Contents 1. GENERAL INFORMATION AND SUMMARY OF THE METHODOLOGY VALIDATION STATUS AND PERFORMANCE CHARACTERISTICS GENERAL COLLABORATIVE TRIAL LIMIT OF DETECTION LIMIT OF QUANTITATION MOLECULAR SPECIFICITY PROCEDURES GENERAL INSTRUCTIONS AND PRECAUTIONS REAL-TIME PCR FOR QUANTITATIVE ANALYSIS OF TC1507 MAIZE General Calibration Real-time PCR set-up DATA ANALYSIS CALCULATION OF RESULTS MATERIALS EQUIPMENT REAGENTS PRIMERS AND PROBES BUFFERS AND SOLUTIONS REFERENCES... 12

62 Protocol TC1507 Community Reference Laboratory 3/17 Document Approval Name / Function Date Signature Marco Mazzara Sector Head Stephane Cordeil Quality Manager Guy Van den Eede B&GMOs Unit Head 21/02/2005 Signed 21/02/2005 Signed 21/02/2005 Signed Address of contact laboratory: European Commission, Joint Research Centre Institute for Health and Consumer Protection (IHCP) Biotechnology and GMOs Unit Community Reference Laboratory Via Fermi 1, Ispra (VA) - Italy

63 Protocol TC1507 Community Reference Laboratory 4/17 1. General information and summary of the methodology This protocol describes an event-specific real-time quantitative TaqMan PCR procedure for the determination of the relative content of event TC1507 DNA to total maize DNA in a sample. The PCR assay has been optimised for use in real-time PCR instruments for plastic reaction vessels. Glass capillaries are not recommended for the buffer composition described in this method. Template DNA extracted by means of suitable methods should be tested for quality ad quantity prior to the use in PCR assay. Tests for the presence of PCR inhibitors (e.g. monitor run, use of DNA spikes) are recommended. For specific detection of event TC1507 genomic DNA, a 58-bp fragment of the recombination region of parts of the construct inserted into the plant genome is amplified using two specific primers. PCR products are measured during each cycle (real-time) by means of a target-specific oligonucleotide probe labelled with two fluorescent dyes: FAM as a reporter dye at its 5 end and TAMRA as a quencher dye at its 3 end. For relative quantitation of event TC1507 DNA, a maize-specific reference system amplifies a 79-bp fragment of HMG (High Mobility Group) gene, a maize endogenous gene, using a pair of HMG gene-specific primers and an HMG gene-specific probe labelled with FAM and TAMRA as described above. The measured fluorescence signal passes a threshold value after a certain number of cycles. This threshold cycle is called the Ct value. For quantitation of the amount of event TC1507 DNA in a test sample, event TC1507 and HMG Ct values are determined for the sample. Standard curves are then used to calculate the relative content of event TC1507 DNA to total maize DNA. 2. Validation status and performance characteristics 2.1 General The method has been optimised for ground maize seed, containing mixtures of genetically modified TC1507 and conventional maize. The reproducibility and trueness of the method was tested through collaborative trial using samples at different GMO contents.

64 Protocol TC1507 Community Reference Laboratory 5/ Collaborative trial The method was validated in a collaborative trial by the Joint Research Centre (JRC) of the European Commission. The study was undertaken with 14 laboratories. Each participant received twelve unknown samples. The samples consisted of DNA mixtures of 0% and 100% TC1507 maize genomic DNA at six GMO levels, between 0.0 % and 5.0 %. Each test sample was analyzed by PCR in three repetitions. The study was designed as a blind duplicate collaborative trial; each laboratory received each level of GM TC1507 in two unknown samples, and the two replicates for each GM level were analyzed on the same PCR plate. A detailed validation report can be found under Limit of detection According the method developer, the absolute LOD of the method is 1.25 copies (8 positives out of 10 replicates). The relative LOD was not assessed in a collaborative trail. The lowest relative concentration of the target sequence included in collaborative trail was 0.1%. 2.4 Limit of quantitation According the method developer, the relative LOQ of the method is 0.08%. The absolute LOQ for the individual systems is 10 copies (TC1507) and 40 copies (HMG). The lowest relative concentration of the target sequence included in collaborative trail was 0.1%. 2.5 Molecular specificity The method utilizes a unique DNA sequence of the recombination region of parts of the construct inserted into the plant genome. The sequence is specific to TC1507 and thus imparts event-specificity to the detection method. The specificity was assessed by Blastsearch on 16/11/2002. No 100% match with other maize GMO sequences was found. The specificity was experimentally tested against DNA extracted from plant materials containing the specific targets of TC1360, Bt176, GA21, NK603, MON810, Bt11, Starlink,

65 Protocol TC1507 Community Reference Laboratory 6/17 T25, MON 863 maize, Roundup Ready soybean, conventional rapeseed, rice and wheat. None of the materials yielded detectable amplification. The target sequence is a single copy sequence in the haploid TC1507 genome. 3. Procedures 3.1 General instructions and precautions All handling of reagents and controls should occur in an ISO 9001:2000 or ISO environment or equivalent. The procedures require experience of working under sterile conditions. Laboratory organization, e.g. flow direction during PCR-setup, should follow the guidelines given by relevant authorities like e.g. ISO, CEN, Codex alimentarius commission. PCR-reagents shall be stored and handled in a separate room and freezer and in equipment where no nucleic acids (with exception of PCR primers or probes) or DNA degrading or modifying enzymes have been handled previously. All handling of PCR reagents and controls requires dedicated equipment especially pipettes. All the equipment used must be sterilized prior to use and any residue of DNA has to be removed. All material used (e.g. vials, containers, pipette tips, etc.) must be suitable for PCR and molecular biology applications. They must be DNase-free, DNA-free, sterile and shall not adsorb protein or DNA. In order to avoid contamination, filter pipette tips protected against aerosol should be used. Use only powder-free gloves and change them frequently. Clean lab-benches and equipment periodically with 10% sodium hypochloride solution (bleach). Pipettes should be checked regularly for precision and calibrated, if necessary. All handling steps - unless specified otherwise - shall be carried out at 0-4 C. In order to avoid repeated freeze/thaw cycles aliquots should be prepared.

66 Protocol TC1507 Community Reference Laboratory 7/ Real-time PCR for quantitative analysis of TC1507 maize General The PCR set-up for the taxon specific target sequence (HMG) and for the GMO (TC1507) target sequence should be carried out in separate vials. Multiplex PCR (using differential fluorescent labels for the probes) has not been tested or validated. The use of maximum 200 ng of template DNA per reaction well is recommended. The method is developed for a total volume of 25 µl per reaction mixture with the reagents as listed in Table 1 and Table Calibration Separate calibration curves with each primer/probe system are generated in the same analytical amplification run. The calibration curves consist of four dilutions of a DNA sample containing 10% TC1507. A series of one to five dilution intervals at a starting concentration of 73,394 maize genome copies may be used (corresponding to 200 ng of DNA with one maize genome assumed to correlate to pg of haploid maize genomic DNA) (Arumuganathan & Earle, 1991). A calibration curve is produced by plotting Ct-values against the logarithm of the target copy number for the calibration points. This can be done e.g. by use of spreadsheet software, e.g. Microsoft Excel, or directly by options available with the sequence detection system software. The copy numbers measured for the unknown sample DNA is obtained by interpolation from the standard curves Real-time PCR set-up 1. Thaw, mix gently and centrifuge the required amount of components needed for the run. Keep thawed reagents at 1-4 C on ice. 2. In two reaction tubes (one for TC1507 system and one for the HMG system) on ice, add the following components (Tables 1 and 2) in the order mentioned below (except DNA) to prepare the master mixes.

67 Protocol TC1507 Community Reference Laboratory 8/17 Table 1. Amplification reaction mixture in the final volume/concentration per reaction well for the reference HMG specific system. Component Buffer 10x (including Rox) Primer MaiJ-F1 Primer mhmg-rev Probe mhmg MgCl 2 25 mm dntps a 10/20 mm AmpliTaq Gold Polymerase Nuclease free water [Template DNA (maximum 200 ng, see and 3.4.2)] Final concentration 1x 300 nm 300 nm 180 nm 4.5 mm 200/400 µm 1U/reaction µl/reaction 2.5 µl µl 0.5 µl - up to 25 µl (5 µl) Total reaction volume: 25 µl a datp (10 mm), dctp (10 mm), dgtp (10 mm), dutp (20 mm) Table 2. Amplification reaction mixture in the final volume/concentration per reaction well for TC1507 specific system. Component Buffer 10x (including Rox) Primer MaiY-F1 Primer MaiY-R3 Probe MaiY-S1 MgCl 2 25 mm dntps a 10/20 mm AmpliTaq Gold Polymerase Nuclease free water [Template DNA (maximum 200 ng, see and 3.4.2)] Final concentration 1x 300 nm 300 nm 150 nm 5.5 mm 200/400 µm 1U/reaction µl/reaction 2.5 µl µl 0.5 µl up to 25 µl (5 µl) Total reaction volume: 25 µl a datp (10 mm), dctp (10 mm), dgtp (10 mm), dutp (20 mm)

68 Protocol TC1507 Community Reference Laboratory 9/17 3. Mix gently and centrifuge briefly. 4. Prepare two reaction tubes (one for the TC1507 and one for the HMG master mix) for each DNA sample to be tested (standard curve samples, unknown samples and control samples). 5. Add to each reaction tube the correct amount of master mix (e.g. 20 x 3 = 60 µl master mix for three PCR repetitions). Add to each tube the correct amount of DNA (e.g. 5 x 3 = 15 µl DNA for three PCR repetitions). Low-speed vortex each tubes at least three times for approx 30 sec. This step is mandatory to reduce the variability among the repetitions of each sample to a minimum. 6. Spin down the tubes in a micro-centrifuge. Aliquot 25 µl in each well. Seal the reaction plate with optical cover or optical caps. Centrifuge the plate at low speed (e.g. approximately 250 x g for 1 minute at 4 C to room temperature) to spin down the reaction mixture. 7. Place the plate into the instrument. 8. Run the PCR with cycling conditions described in Table 3: Table 3. Reaction conditions. Step Stage T C Time (sec) Acquisition Cycles 1 Initial denaturation 95 C 600 No 1x 2a Denaturation 95 o C 15 No 2b Amplification Annealing & Extension 60 o C 60 Measure 45x 3.3 Data analysis Subsequent to the real-time PCR, analyse the run following the procedure below: a) Set the threshold: display the amplification curves of one system (e.g. TC1507) in logarithmic mode. Locate the threshold line in the area where the amplification profiles are parallel (exponential phase of PCR) and where there is no fork effect between repetitions of the same sample. Press the update button to ensure changes affect Ct values. Switch to the linear view mode by clicking on the Y axis of the amplification plot, and check that the threshold previously set falls within the geometric phase of the curves.

69 Protocol TC1507 Community Reference Laboratory 10/17 b) Set the baseline: determine the cycle number at which the threshold line crosses the first amplification curve and set the baseline three cycles before that value (e.g. earliest Ct = 25, set the baseline crossing at Ct = 25 3 = 22). c) Save the settings d) Repeat the procedure described in a) and b) on the amplification plots of the other system (e.g. HMG system). e) Save the settings and export all the data into an Excel file for further calculations. 3.4 Calculation of results After having defined a threshold value within the logarithmic phase of amplification as described above, the instruments software calculated the Ct-values for each reaction. The standard curves are generated both for the HMG and TC1507 specific systems by plotting the Ct-values measured for the calibration points against the logarithm of the DNA copy numbers, and by fitting a linear regression line into these data. Thereafter, the standard curves are used to estimate the copy numbers in the unknown sample DNA by interpolation from the standard curves. For the determination of the amount of TC1507 DNA in the unknown sample, the TC1507 copy number is divided by the copy number of the maize reference gene (HMG) and multiplied by 100 to obtain the percentage value (GM% = TC1507/HMG * 100). 4. Materials 4.1 Equipment Real-time PCR instrument for plastic reaction vessels (glass capillaries are not recommended for the described buffer composition) Plastic reaction vessels suitable for real-time PCR instrument (enabling undisturbed fluorescence detection) Software for evaluating data after standard curve method (mostly integrated in the software of the real-time PCR instrument) Microcentrifuge

70 Protocol TC1507 Community Reference Laboratory 11/17 Micropipettes Vortex Rack for reaction tubes 1.5/2.0 ml tubes 4.2 Reagents (equivalents may be substituted) TRIS ph=8.0: Tris[hydroxymethyl] aminomethane hydrochloride (Molecular Biology grade) KOAc (SIGMA Part No P1190) Gelatine (VWR Part No ) Tween 20 (SIGMA Part No P ML) Glycerol (SIGMA Part No P ML) Rox (Applied Biosystems Part No ) datp (GeneCraft Part No GC ) dctp (GeneCraft Part No GC ) dgtp (GeneCraft Part No GC ) dutp (GeneCraft Part No GC ) MgCl 2 (SIGMA Part No M1028-1ML) Ampli Taq Gold (Applied Biosystems Part No N ) 4.3 Primers and Probes Name Oligonucleotide DNA Sequence (5 to 3 ) TC1507 target sequence MaiY-F1 TAG TCT TCG GCC AGA ATG G MaiY-R3 CTT TGC CAA GAT CAA GCG MaiY-S1 6-FAM-TAA CTC AAG GCC CTC ACT CCG-TAMRA Reference gene HMG target sequence MaiJ-F2 TTG GAC TAG AAA TCT CGT GCT GA mhmg-rev GCT ACA TAG GGA GCC TTG TCC T Mhmg-probe 6-FAM-CAA TCC ACA CAA ACG CAC GCG TA-TAMRA

71 Protocol TC1507 Community Reference Laboratory 12/17 5. Buffers and Solutions The following describes the preparation of the buffers used in this procedure. Volume may be scaled as needed. Equivalent reagent may be substituted. Preparation of the 10x Buffer a) Mix the following chemicals at the final concentration indicated and adjust the buffer to ph = 8.0 Component Final concentration Tris ph = M KOAc 0.5 M Gelatine 0.5% Tween % Glycerol 0.8% Rox 0.2 µl/reaction Water - 6. References Arumuganathan, K., Earle, E.D. (1991). Nuclear content of some important plant species. Plant Mol Biol Reporter 9,

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88 Liliana Irene Sánchez Cortés MCP - Maestría en Ciencias en Sist. Calidad y Productividad Lugar de nacimiento: León, Guanajuato, México Dirección actual: CIRCUITO SERENIDAD #52, VILLASERENIDAD, Bahia de Banderas, Nayarit, México. Teléfono: Casa: (322) Celular: (322) Correo electrónico : [email protected] Educación1 MCP - Maestría en Ciencias en Sist. Calidad y Productividad ITESM Rectoría Universidad Virtual ( ) México Master en Paisaje UNIVERSIDAD POLITECNICA DE MADRID ( ) España Intercambio, tesis y disertación: Diseño de proyecto de parque público para la facultad de agronomía de la Universidad Politécnica de Madrid (Análisis de impacto ambiental, costos, plan de negocio, diseño, etc.) IAP - Ingeniero Agrónomo en ITESM Campus Queretaro ( ) Producción Área de concentración: Especialidad en Acuacultura Intercambio, tesis y disertación: - Intercambio Universidad Pontificia de Comillas Madrid, España. Curso de Valores Éticos para la profesión - ITESM Sede Guayaquil, Ecuador. Semestre de Especialidad en Acuacultura y Finanzas. - Residencia de Verano en Grupo Pulsar, Comitán, Chiapas. Invernaderos de Producción de Semilla Híbrida de Hortalizas. - ITESM Campus Querétaro. Tesis, Plan de negocios y análisis financiero para proyecto de produccion de ornamentales bajo invernadero. Áreas de experiencia Agricultura / Ganadería 5 Años Calidad 4 Años Seguridad 4 Años Manufactura, producción y operaciones - 3 Años Equipos de alto desempeño Manufactura, producción y operaciones - Evaluación / Control de procesos Compras y logística - Importación / Exportación 3 Años Manufactura, producción y operaciones - Mantenimiento 3 Años 4 Años Ecología 3 Años Experiencia profesional PIONEER HI-BRED INTERNATIONAL Agrícola PUERTO VALLARTA,México SENIOR RESEARCH SUPERVISOR ( Actual) Principales responsabilidades: Coordinar y supervisar los departamentos de investigación de maíz y operaciones: - Planeación, revisión y control de Presupuestos y Capital para los departamentos en conjunto con el área administrativa y el grupo de liderazgo - Dirigir y administrar el sistema de aseguramiento de calidad de la empresa mediante la planeación e implementación de proyectos de mejora continua - Supervisar la evaluación y control de procesos de Seed Handling

89 - Administrar recursos y sistemas de procesamiento para dar cumplimiento a procesos, instrucciones y sistemas de trabajo organizacionales y de calidad - Supervisar la coordinación de tramites de regulación para importaciones, exportaciones y ambiental - Coordinador SHE (Safety, Health and Environment) - Lean practicioner Principales logros: Liderar un grupo de ingenieros y técnicos en el área de operaciones y maíz para lograr los objetivos de calidad, lograr las metas de los programas de cultivos y todas las actividades requeridas para cumplir las necesidades internas y de los clientes. Crear planes estratégicos y operativos para las mejoras de calidad de la compañía. Lograr la implementación del sistema ISO9001:2000 y lograr la certificación. Participar activamente dentro de los programas y grupos de la compañía para trabajar sobre metas específicas entre localidades a nivel internacional. VIVEROS GDV Florería / Viveros León,México INGENIERO DE PROYECTOS ( ) Principales responsabilidades: COORDINACIÓN DE PROYECTOS URBANOS, PARQUES INDUSTRIALES, ENTRE OTROS. ANÁLISIS DE COSTOS E IMPACTO AMBIENTAL. ACCESORIOS Y SISTEMAS HIDRÁULICOS SA DE CV Agrícola AGUASCALIENTES,México INGENIERO DE PROYECTOS ( ) Principales responsabilidades: DISEÑAR Y DIRIGIR LAS ACTIVIDADES DE DISEÑO DE SISTEMAS DE RIEGO AGRÍCOLA, TRABAJO CON EL GOBIERNO EN PROYECTOS DE APOYO Y SUBSIDIOS. MANEJO DE EQUIPO DE PERSONAL DE CAMPO E INSTALADORES. Principales logros: PARTICIPAR ACTIVAMENTE CON EL GOBIERNO DEL ESTADO DE AGUASCALIENTES EN PROYECTOS DE SISTEMAS DE RIEGO. APLICACION DE HABILIDADES Y CONOCIMIENTOS TÉCNICOS EN CAMPO. MANEJO DE UN PERSONAL, VIAJANDO CONSTANTEMENTE. Paquetes computacionales Software MS Office - Avanzado Access - Intermedio Lotus Smartsuite - Intermedio PhotoShop - Intermedio SAP - Intermedio AutoCad - Intermedio CorelDraw - Intermedio Adobe After Effects - Básico Idiomas Español - Avanzado Inglés - Avanzado Francés - Básico Experiencia internacional: Tiempo en el extranjero: Países en los que realizó los proyectos más importantes: Proyectos realizados: 2 años España MAESTRIA EN PAISAJISMO Y TRABAJO DE MEDIO TIEMPO EN EL ÁREA DE DESARROLLO DE PROYECTOS, PRESUPUESTOS, ETC.

90 ENRIQUE CAMPOS WHITE Los Robles # 21 Frac. Campestre Tres Pasos Xalapa, Veracruz, México Casa: (228) Cell: (55) personal: [email protected] PERFIL PROFESSIONAL Ejecutivo en Comercializar y Desarrollar Tecnología con nueve años de experiencia probada en cultivos algodón y soya Modificados Genéticamente (GMO) así como en el mercado de maíz, con una formación en el manejo de herbicidas. Líder Participativo en el desarrollo e implementación de Programas Comerciales y de Mercado, Propuestas de Valor, programas de entrenamiento en cultivos de maíz, algodón y soya así como de herbicidas, con un rol activo en la generación de demanda y valor agregado. OBJETIVO Obtener una posición ejecutiva en una Compañía Agrícola líder en su ramo, donde pudiera contribuir con mis conocimientos y habilidades técnicos-comerciales, valorando la importancia del factor humano como el mayor y principal activo y teniendo una visión corporativo a mediano-largo plazo. RESUMEN DE CAPACIDADES Poseo la capacidad de: Producir credibilidad y confianza por mis habilidades técnicas y comerciales; trabaja en equipo; administrar de manera eficiente recursos limitados para proyectos específicos; motivar a la gente bajo condiciones limitadas; lograr crear equipos de trabajo comprometidos y enfocados a resultados; motivar, desarrollar y direccionar a la gente en sus trabajos e identificando a la mejor gente para cada trabajo; enfocarme y orientar las acciones a los resultados; trabajar bajo presión; enseñar a otros como hacer las cosas; generar o adaptar equipos y tecnología a diferentes necesidades; ser flexible y adaptarme a nuevos cambios; identificar problemas potenciales y desarrollar información necesaria para desarrollar y evaluar opciones e implementar soluciones; sopesar costos y beneficios de alguna acción potencial. ANTECEDENTES LABORALES Pioneer, A DuPont Company Tel. (33) División Investigación: Gerente de Investigación Enero 2008 a la fecha Arysta LifeScience Corporation Tel. (55) División Comercial: Gerente de Marketing Mayo a Diciembre 2007

91 Semillas Cristiani Burkard Tel. (961) División Comercial: Gerente de Ventas Veracruz Septiembre 2006 a abril Monsanto Company Tel. (55) y (33) Negocio de Biotecnología-Agroquímicos: Gerente Nacional de Tecnología para México Agosto 2001 a Diciembre Negocio de Agroquímicos: Gerente de Marketing para cultivos perennes en México. Marzo 2000 a Septiembre Gerente de Tecnología para el Sureste Mexicano. Diciembre 1998 a Febrero Gerente Local de Ventas y Marketing para Chiapas. Enero 1995 a Noviembre ITESM (Campus Monterrey) Centro de Planeación Estratégica: Asistente de Investigación Agosto 1993 a Diciembre SAGARPA (Delegación Aguascalientes) Campañas Fitosanitarias: Jefe de Campañas fitosanitarias Junio a Diciembre Banca Serfín División Jurídica, Cd. de Mexico: Perito Agrícolas y Ganadero Enero 1992 a Abril Corporaciones G.A.L. (Aguascalientes) División Agrícola y Ganadera Director de Producción Febrero 1991 a Diciembre Corporaciones Valle de Pabellón de Arteaga División Agrícola Gerente General Marzo 1989 a Diciembre Mesa Grande Sociedad de Producción Rural y de R.I. División Agrícola Gerente Técnico Junio 1986 a Febrero PRINCIPALES LOGROS Y RESPONSABILIDADES Pioneer, A DuPont Company Gerente de Investigación: Reporte a la Dirección General de Investigación. Liderar las operaciones del NRSP in Puerto Vallarta, dando el soporte necesario a los grupos de maiz- MPD, soya, sorgo-cpd. Planear y conducir programas e investigación. Identificar y conocer las necesidades de los clientes internos y externos. Lederear los proceses administrativos, de reclutamiento de personal y manejo de personal. Proveer los recursos necesarios a las diferentes áreas de mantenimiento, así como desarrollar los programas de infraestructura, logística de seguridad, regulación e ISO Responder por las actividades de manejo de semilla Arysta LifeScience Gerente de Marketing: Reporte dual a la Dirección General y a la Gerencia Comercial.

92 Tengo la responsabilidad de elaborar el Plan de mercadeo para los diferentes productos del portafolio. Como parte del plan de mercadeo tengo que desarrollar la estrategia de venta del portafolio en cada uno de los diferentes cultivos. De acuerdo al conocimiento de los productos y de los diferentes mercado debo elaborar el Posicionamiento o reposicionamiento de los diferentes productos por cultivo. Debo también buscar la rentabilidad por cada uno de los productos de acuerdo a las situaciones específicas de cada mercadeo y en cada región. Como parte importante de esta posición debo buscar nuevas oportunidades e innovar diferentes formas de uso de los productos en los diferentes cultivos donde participamos actualmente. Generar información para la Fuerza de ventas con respecto al mercado, competencia de otras compañías y de moléculas genéricas, oportunidades de nuevos negocios, etc. Coordinar el desarrollo de nuevos productos junto con el Departamento de Desarrollo de acuerdo a cada mercado y cada cultivo donde participamos o tenemos la oportunidad de hacerlo. Coordinar junto con la Fuerza de ventas y la Gerencia Comercial las propuestas de ventas de acuerdo a la oportunidad del cultivo y la situación específica del mercado a mediano y largo plazo. Semillas Cristiani Burkard Coordinador de Ventas Veracruz: Reporte a la Gerencia General. Lideré un grupo de tres Agrónomos empleados temporales, donde siempre se busco resaltar valores como trabajo en equipo, dedicación, lealtad, perseverancia, honestidad y un alto compromiso hacia la Compañía. Inicie la estructuración de una nueva red de distribución que fuera capaz de posicionar los productos de Cristiani como la empresa numero uno en la comercialización de semilla de maíz, sorgo y pastos en Veracruz antes del Identifique y comencé el desarrollo de distribuidores más enfocados para el mercado de semillas con perfil empresarial. Identifique el Potencial, definir y desarrolle la estrategia comercial para lograr una participación del mercado de maíz en Veracruz del 12%, en sorgo del 5% y en pastos del 10% durante el ciclo agrícola Negocie y firme convenios de venta y distribución con la nueva red, así como realicé firma de convenios con la cadena de sub-distribución del territorio a mi cargo. Elabore junto con el distribuidor los programas de trabajo y al mismo tiempo asegurar la correcta implementación y enfoque de los mismos. Desarrolle una compañía de mercadeo que permitiera logara la penetración y participación de mercado deseado y posicionar la marca como los maíces tropicales de mejor calidad y rendimiento en Veracruz. Seleccione, capacite y supervise grupos de Promotores técnicos-comerciales, bajo un ambiente participativo y altamente competitivo. Monsanto Company Como Líder de Tecnología en Biotecnología y Agroquímicos: Reporte dual a la Dirección de Marketing y a la Dirección de Tecnología. Lideré un grupo de doce Agrónomos donde siempre se busco resaltar valores como trabajo en equipo, dedicación, lealtad, perseverancia, honestidad y un alto compromiso hacia la Compañía. Con la información generada por el Departamento bajo mi responsabilidad tuve siempre roles activos y determinantes en la planeación, desarrollo e implementación de los programas comerciales y de mercadeo. Tenía la corresponsabilidad junto con el Gerente Comercial en planear y ejecutar los programas comerciales para algodón y soya, así como soportar con la información necesaria para determinar los precios de comercialización de nuestras tecnologías. Tenía la corresponsabilidad junto con el Gerente de Mercadeo en desarrollar los posicionamientos en el uso de nuestras tecnologías en los cultivos modificados genéticamente, así como el tener un rol activo en la implementación de estos programas. Tenía también la corresponsabilidad de supervisar, retroalimentar, informar y motivar a los representantes de ventas y promotores de campo con la finalidad de asegurar los usos correctos

93 de las tecnologías cumpliendo con los reglamentos regulatorios, políticas comerciales y posicionamientos de mercadeo. Formaba parte del Equipo Regulatorio de la Compañía, teniendo como responsabilidad básica el generar la información de campo requerida por las Autoridades Regulatorias Federales y Estatales. Nuestra equipo tenia la responsabilidad en la elaboración de informes del comportamiento de estas tecnologías en campo, resaltando los impactos económicos, agronómicos y del medio ambiente a estas tecnologías en las regiones donde se evaluaban y/o comercializaban estos cultivos, para las autoridades regulatorios federales y estatales. Mantenía una estrecha relación con las autoridades Regulatorias Estatales informando y mostrando en campo y con los productores los benéficos de estas tecnologías en las diferentes zonas. Tenia participación activo en diferentes foros en las distintas regiones donde se siembran estos cultivos. Lidere el programa de stewardship para los cultivos modificados genéticamente en México y como principal preocupación la adecuada introducción de Bollgard II y Solución Faena Flex a partir del ciclo Nuestro equipo determino los parámetros y desarrollo la(s) Propuesta(s) de Valor para alfalfa Solución Faena en el Norte de México, con un potencial de Mercado estimado en 200 k ha. Iniciamos el desarrollo de las Propuestas de Valor de las tecnologías en algodón Bollgard II y Solución Faena Flex, con la finalidad de sustituir tecnologías existentes en México como Bollgard y Solución Faena. Lideré el programa de selección de semillas de algodón de la marca Stoneville, propiedad de Monsanto, buscando siempre aquellas variedades con mejor adaptación agronómica tanto de materiales modificados genéticamente como convencionales en las diferentes regiones de Mexicali, Sonora, Chihuahua y Tamaulipas. Coordine el programa conjunto Monsanto-D&PL de selección de semillas de algodón, buscando aquellas variedades con mejor adaptación agronómica, así como aquellos materiales que mejor se comportaran al Sistema Solución Faena-Bollgard en las diferentes regiones de Mexicali, Sonora y Chihuahua, así como necesidades comerciales y de rentabilidad. Lideré el programa de selección de semillas de soya, buscando aquellas variedades con mejor adaptación agronómica y que mejor encajaran a las propuestas de valor en las regiones de Chiapas, Tamaulipas y Sinaloa. Pudimos entender el potencial de Solución Faena Algodón y Solución Faena Soya, no solamente como una tecnología para el control de maleza sino también como un Sistema integral de manejo de estos cultivos, con un potencial en ambos cultivos en Mexico de 200 k ha. Desarrolle una metodología para el uso de Solución Faena Soya en rotación con maíz en Sinaloa con un mercado estimado in 400 k ha. Desarrollamos las Propuestas de Valor para Solución Faena Soya y Solución Faena algodón, participando en el desarrollo de un nuevo negocio en Mexico con un valor estimado de $6 M Usd. Desarrolle nuevas propuestas de valor para algunos cultivos perennes como agave, logrando con esto recuperar y crecer Mercado en compañías como Casa Cuervo y Tequila Sauza y ajuste propuestas añejas logrando nuevos nichos en mercados maduros en cultivos como café y cítricos, logrando crecimientos adicionales de 30% y 15% respectivamente. Por estas acciones nuestro equipo fue reconocido por estas compañías como Casa Cuervo y Tequila Sauza, al reducir los costos de control de maleza en mas de 35% (equivalente a ahorros de $0.3 M Usd por año). Desarrolle nuevos productos a base de Glifosato como Faena con Transorb, Faena Ultra y Faena Fuerte con Transorb y otro tipo de herbicidas como Sempra (Halosulfuron) y Defensa (Picloran). Diseñe un aspersor para aplicaciones seguras de Glifosato en agave; posteriormente fue comercializada por compañías como Swissmex y FMA. Desarrolle programas de manejo integral de maleza (MIM) para cítricos en Veracruz, Colima y Apatzingan, así como para café en Tapachula, Coatepec y Xicotepec y para banano en Tapachula y Tecomán. Frecuentemente participe en seminarios técnicos, simposios, entrenamientos para distribuidores y personal de la Compañía, siempre con una alta calidad en la información generada. Para poder lograr estas metas tuve que cambiar de residencia como Xalapa y Guadalajara. Como Gerente de Marketing para cultivos perennes en México: Identifiqué el Potencial y definí la estrategia comercial para los Cultivos Perennes en México, con un valor actual de ventas por $24.7 M Usd.

94 Diseñé e implementé campañas publicitarias y promociónales logrando enfocar nuestro portafolio de productos y servicios en los diversos segmentos del Mercado en específico los mercados de alto valor como café, cítricos y agave. Desarrollé diversos segmentos de mercado logrando penetrar nuevos productos para la compañía, así como diversifiqué y consolidé el portafolio en mercados maduros logrando retener o recapturar segmentos con baja o nula participación. Elaboré y desarrollé estratégica y comercialmente diversos esquemas Vs. Genéricos en los segmentos más hostiles / baja rentabilidad para la compañía. Estructuré la logística de diversas campañas de lanzamiento de herbicidas como Faena Ultra, Faena Full y Faena con Transorb ; logrando siempre los resultados esperados internamente (dentro de los tiempos y presupuesto planeado) y externamente (desarrollo y penetración de las marcas). Realicé programas de Co-promoción buscando sinergizar el desarrollo y transferencia de tecnología al mercado para nuestros productos y/o servicios, con empresas afines en la industria, entre otras Sanz Hermanos y FMA. Brindé mi total disponibilidad radicando en la Cd. de Xalapa para capitalizar lo anterior. Como Gerente Local de Ventas y Marketing: Demostré una buena capacidad de Planeación y excelente Ejecución para estructurar la cadena de distribución en cada una de las regiones a mi cargo. Teniendo bajo mi responsabilidad el principal distribuidor del Sureste de México durante 3 años consecutivos. Mantuve una Red de distribución altamente motivada, enfocada, leal a la marca y con un alto conocimiento del desempeño de nuestros productos de acuerdo a cada segmento de mercado. Elaborar junto con el distribuidor los programas de trabajo y al mismo tiempo asegurar la correcta implementación y enfoque de los mismos. Direccioné los esfuerzos comerciales de la compañía para capturar y retener los principales mercados potenciales del Sureste. Logré consistentemente los presupuestos de venta establecidos en el mediano y largo plazo, dí claridad al potencial real de la zona para continuar el desarrollo comercial de la misma. Fui capaz de desarrollar el mercado en mas de un 300% en 3 años, yendo de $1.9M Usd a $4.2M Usd. Seleccioné, capacité y supervisé grupos de Representantes y Asesores técnicos y de ventas, bajo un ambiente participativo y altamente competitivo. Negocié y logré la firma de los convenios de venta y distribución con las principales cuentas mayoristas del Sureste. Además realicé la firma de convenios con la cadena de sub-distribución del territorio a mi cargo. Para lograr lo anterior brindé mi total disponibilidad para radicar en la ciudad de Tapachula, Chis. RECONOCIMIENTOS Monsanto World Wide Master Sales Award en 1996 (Reconocimiento Mundial de ventas). Achievement Award en 1995 y 1997 (Reconocimiento Nacional de ventas). Fast Recognition en 2002 y 2004 (Por contribuciones Técnicas en el negocio de Biotecnología). Purdue University (Indiana, US) Especialización en Herbicidas (1999). FORMACION ACADEMICA I.T.E.S.M. (Campus Monterrey) Maestría en Ciencias Especializado en Fitosanidad ( ). Universidad Autonoma de Aguascalientes Ingeniería en Agronomía con especialidad en Fruticultura ( ). Idiomas: Español e Ingles (TOFEL: 604).

95 CURSOS Y SEMINARIOS RELEVANTES Entendimiento del Cliente / México, D.F. Brain Effects FX Habilidades Personales / México D.F. MERCER. Human Resources Consulting Ventas Efectivas con el uso de la Psicología / México D.F. Dimensional Solutions Team Work, Developing Human Resources Potential For Businesses Cornellius and Associates Leadership through People Skills / St.Louis, Mo. US. Negociación Efectiva Socratic Selling / Mexico, D.F. Los 7 Hábitos de la Gente Altamente Eficiente / México, D.F. Programa de Ventas Dimensionales / Toluca, Edo.Méx. Habilidades Financieras y de Marketing / México D.F. Monsanto Entrenamiento. AMIPFAC. Uso Seguro de Agroquimicos / Veracruz, BUMA. Marketing Aplicada / Xalapa COPARMEX Comunicación Abierta / Cuernavaca, Mor. Integración y Motivación de Equipo Comercial / Avándaro, Edo.Méx. Entrenamiento de Microsoft Office, Word, EXCEL, Power Point, Harvard Graphics y Lotus cc mail / Mexico D.F. Monsanto. INFORMACIÓN PERSONAL Estado civil: Casado con 3 hijos de 16, 12 y 6 años. Fecha de Nacimiento: 18 de marzo de Nacionalidad: Mexicano Elaborado en Julio de 2007.

96 ROBERTO CAMBEROS AGUAS Nicolás Bravo # 179 Colonia Independencia Puerto Vallarta, Jalisco, México C.P Teléfono: (322) (Casa) Móvil: (322) (329) y (Oficina), [email protected] [email protected] INFORMACION PERSONAL Lugar de Nacimiento: Guadalajara, Jal. México Fecha de Nacimiento: 13 de Mayo de 1968 Estado Civil: Casado Dependientes: 3 (Esposa e hijas) IITERES PROFESIONAL. Colaborar en los programas de mejoramiento genético de maíz a nivel mundial dentro de Pioneer, así como en la producción se semilla para investigación, aplicación de nuevas tecnologías tales como organismos genéticamente modificados y dobles haploides. EXPERIENCIA PROFESIONAL: Marzo 2008 a la Fecha: Cargo: Corn Senior Research Associated-NRSP Compañía: PHI Servicios S.A. de C.V. Camino Viejo a Valle de Banderas Km. 3 # 19, Tapachula Nayarit, México. Princilales Responsabilidades: Liderear y coordinar la operación del departamento de maíz, supervisando además que se cumplan resultados de acuerdo a los estándares establecidos por la compañía. Planeación de actividades y asignación de campos de acuerdo a fechas de siembra. Planear parte del presupuesto que se asignará a la operación del programa de maíz de acuerdo a carga de trabajo y personal de campo a contratar.

97 Enero 2005 a Febrero 2008: Cargo: Corn research associate-nrsp. Compañía: PHI Servicios S.A. de C.V. Camino Viejo a Valle de Banderas Km. 3 # 19, Tapachula Nayarit, México. Principales Responsabilidades: Llevar a cabo todas las actividades del programa de maíz incluyendo (siembras, polinizaciones, desespigues, cosecha, toma de datos, etc.) para lograr los targets de semilla requeridos así como buena calidad y cantidades de semilla. Manejo de PRISM (Abrir localidades, publicar localidades, mapeos, libros de campo, etiquetas de campo, requisición de semilla tanto para lotes aislados como viveros etc.) Coordinar y supervisor la preparación de semilla, planeación de las fechas de siembra de acuerdo al calendario establecido por el equipo de maíz en USA. Coordinar y supervisor el equipo de polinizadores de campo (cerca de 130 personas) durante la temporada de polinizaciones de los viveros Coordinar y dar seguimiento a experimentos especiales establecidos en la estación experimental de Puerto Vallarta: o Observación de líneas o Estudio de la sensibilidad al secado o Colección de datos (floración, tamaño de espiga, humedad de cosecha, % capa negra, % línea de leche, etc.) así como otros traits requeridos por los mejoradores. o Participación como coordinador del procedimiento de calibración en el equipo de ISO 9001 desde Octubre 1997 a Diciembre 2004: Antes de la integración del grupo de operaciones en Puerto Vallarta tuve otras responsabilidades tales como: Todas las actividades relacionadas a importación y exportación de semilla, manejo diario de actividades regulatorias fitosanitarias (Certificados fitosanitarios para importación y exportación de semilla, y dar seguimiento a los procesos cuarentenarios establecidos por leyes regulatorias de México) Coordinar actividades de desgrane y empaque de semilla de maíz. Doblamiento, desarrollo e incremento de semilla doble haploide.

98 Octubre 1996, a Octubre 1997: Cargo: Supervisor de Campo. Compañia: Semillas Híbridas S.A. de C.V. (Dekalb Genetics) Principales Responsabilidades: Durante este periodo de tiempo trabajé para, Dekalb Genetics dentro del departamento de producción. Durante este periodo mis principales responsabilidades fueron la producción de semilla, contactando productores, contratando tierras para la producción de semilla comercial y parental. Dando seguimiento a todas las localidades durante su desarrollo, controlando además la presencia de plagas, malezas y enfermedades. De igual manera yo era responsable de contratar y coordinar las cuadrillas de desespigadores (cerca de 150 personas en total) December 1992 to December 1995: Cargo: Polinizador temporal (lider de grupo). Compañia: Híbridos Pioneer de México S.A. de C.V. Camino Viejo a Valle de Banderas Km 3 # 19, Tapachula Nayarit, México. Durante 4 temporadas de polinizaciones trabajé como empleado eventual apoyando polinizaciones en maíz desespigues y otras actividades en aislamientos y viveros de producción de semilla para investigación. EDUCACION: Ingeniero Agrónomo: Septiembre 992 a Agosto 1996; Centro Universitario de Ciencias Biológicas y Agropecuarias, Universidad de Guadalajara, Las agujas, Mpio. de Zapopan, Jal. México.

99 Certificado recibido: Titulo de Ingeniero Agrónomo Fitotcnista. HABILIDADES: Competente en el manejo de varios programas de computación (Excel, Word, Power point, bartender, Photoshop, PRISM etc.) Conocimiento en la operación de SQUID (Seed Quality Improvement Device), operación de SSTS (Seed Sample Tracking and Shipment), y software de colección de datos. Idiomas: Español (lengua nativa) Inglish (Intermedio alto) ENTRENAMIENTOS Y CURSOS: Junio, Julio y Agosto del 2004: Entrenamiento en la metodología de Six sigma (Producción de semilla doble haploide en dos diferentes condiciones ambientales.) Junio-Julio, 2003: Entrenamiento en Tecnología de dobles haploides en la estación experimental de Waimea, HI. El entrenamiento consistió en el aprendizaje de doblamiento con técnicas de laboratorio, transplante a campo, polinización y cosecha de material doble haploide. Junio del 2002, Entrenamiento de PRISM por 5 días en Johnston, IA, USA.

100 Publicaciones. Sandoval I. E., L. J. Arellano R. L., R. O. Garay V., R. Camberos A. Evaluación de 25 híbridos de sorgo (Sorghum bicolor L. Moench) en Zapopan, Jalisco. Universidad de Guadalajara, Facultad de Agronomía. Resultados y avances de investigación. November, González L. S., E. Sandoval I., L. Varela R., R. Camberos A. Efectos de productos hormonales sobre la viabilidad y emergencia en semillas de maíz (Zea mays L.). Universidad de Guadalajara, Centro Universitario de Ciencia Biológicas y Agropecuarias. Resultados y avances en Investigación. May, 1994.

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104 INTERNATIONAL STANDARD ISO 9001 Fourth edition Quality management systems Requirements Systèmes de management de la qualité Exigences Reference number ISO 9001:2008(E) ISO 2008

105 ISO 9001:2008(E) PDF disclaimer This PDF file may contain embedded typefaces. In accordance with Adobe's licensing policy, this file may be printed or viewed but shall not be edited unless the typefaces which are embedded are licensed to and installed on the computer performing the editing. In downloading this file, parties accept therein the responsibility of not infringing Adobe's licensing policy. The ISO Central Secretariat accepts no liability in this area. Adobe is a trademark of Adobe Systems Incorporated. Details of the software products used to create this PDF file can be found in the General Info relative to the file; the PDF-creation parameters were optimized for printing. Every care has been taken to ensure that the file is suitable for use by ISO member bodies. In the unlikely event that a problem relating to it is found, please inform the Central Secretariat at the address given below. ISO 2008 COPYRIGHT PROTECTED DOCUMENT All rights reserved. Unless otherwise specified, no part of this publication may be reproduced or utilized in any form or by any means, electronic or mechanical, including photocopying and microfilm, without permission in writing from either ISO at the address below or ISO's member body in the country of the requester. ISO copyright office Case postale 56 CH-1211 Geneva 20 Tel Fax [email protected] Web Published in Switzerland ii ISO 2008 All rights reserved

106 ISO 9001:2008(E) Contents Page 1 Scope General Application Normative references Terms and definitions Quality management system General requirements Documentation requirements Management responsibility Management commitment Customer focus Quality policy Planning Responsibility, authority and communication Management review Resource management Provision of resources Human resources Infrastructure Work environment Product realization Planning of product realization Customer-related processes Design and development Purchasing Production and service provision Control of monitoring and measuring equipment Measurement, analysis and improvement General Monitoring and measurement Control of nonconforming product Analysis of data Improvement Annex A (informative) Correspondence between ISO 9001:2008 and ISO 14001: Annex B (informative) Changes between ISO 9001:2000 and ISO 9001: Bibliography ISO 2008 All rights reserved iii

107 ISO 9001:2008(E) Foreword ISO (the International Organization for Standardization) is a worldwide federation of national standards bodies (ISO member bodies). The work of preparing International Standards is normally carried out through ISO technical committees. Each member body interested in a subject for which a technical committee has been established has the right to be represented on that committee. International organizations, governmental and non-governmental, in liaison with ISO, also take part in the work. ISO collaborates closely with the International Electrotechnical Commission (IEC) on all matters of electrotechnical standardization. International Standards are drafted in accordance with the rules given in the ISO/IEC Directives, Part 2. The main task of technical committees is to prepare International Standards. Draft International Standards adopted by the technical committees are circulated to the member bodies for voting. Publication as an International Standard requires approval by at least 75 % of the member bodies casting a vote. Attention is drawn to the possibility that some of the elements of this document may be the subject of patent rights. ISO shall not be held responsible for identifying any or all such patent rights. ISO 9001 was prepared by Technical Committee ISO/TC 176, Quality management and quality assurance, Subcommittee SC 2, Quality systems. This fourth edition cancels and replaces the third edition (ISO 9001:2000), which has been amended to clarify points in the text and to enhance compatibility with ISO 14001:2004. Details of the changes between the third edition and this fourth edition are given in Annex B. iv ISO 2008 All rights reserved

108 ISO 9001:2008(E) Introduction 0.1 General The adoption of a quality management system should be a strategic decision of an organization. The design and implementation of an organization's quality management system is influenced by a) its organizational environment, changes in that environment, and the risks associated with that environment, b) its varying needs, c) its particular objectives, d) the products it provides, e) the processes it employs, f) its size and organizational structure. It is not the intent of this International Standard to imply uniformity in the structure of quality management systems or uniformity of documentation. The quality management system requirements specified in this International Standard are complementary to requirements for products. Information marked NOTE is for guidance in understanding or clarifying the associated requirement. This International Standard can be used by internal and external parties, including certification bodies, to assess the organization's ability to meet customer, statutory and regulatory requirements applicable to the product, and the organization's own requirements. The quality management principles stated in ISO 9000 and ISO 9004 have been taken into consideration during the development of this International Standard. 0.2 Process approach This International Standard promotes the adoption of a process approach when developing, implementing and improving the effectiveness of a quality management system, to enhance customer satisfaction by meeting customer requirements. For an organization to function effectively, it has to determine and manage numerous linked activities. An activity or set of activities using resources, and managed in order to enable the transformation of inputs into outputs, can be considered as a process. Often the output from one process directly forms the input to the next. The application of a system of processes within an organization, together with the identification and interactions of these processes, and their management to produce the desired outcome, can be referred to as the process approach. An advantage of the process approach is the ongoing control that it provides over the linkage between the individual processes within the system of processes, as well as over their combination and interaction. When used within a quality management system, such an approach emphasizes the importance of a) understanding and meeting requirements, b) the need to consider processes in terms of added value, ISO 2008 All rights reserved v

109 ISO 9001:2008(E) c) obtaining results of process performance and effectiveness, and d) continual improvement of processes based on objective measurement. The model of a process-based quality management system shown in Figure 1 illustrates the process linkages presented in Clauses 4 to 8. This illustration shows that customers play a significant role in defining requirements as inputs. Monitoring of customer satisfaction requires the evaluation of information relating to customer perception as to whether the organization has met the customer requirements. The model shown in Figure 1 covers all the requirements of this International Standard, but does not show processes at a detailed level. NOTE In addition, the methodology known as Plan-Do-Check-Act (PDCA) can be applied to all processes. PDCA can be briefly described as follows. Plan: establish the objectives and processes necessary to deliver results in accordance with customer requirements and the organization's policies. Do: implement the processes. Check: monitor and measure processes and product against policies, objectives and requirements for the product and report the results. Act: take actions to continually improve process performance. Figure 1 Model of a process-based quality management system vi ISO 2008 All rights reserved

110 ISO 9001:2008(E) 0.3 Relationship with ISO 9004 ISO 9001 and ISO 9004 are quality management system standards which have been designed to complement each other, but can also be used independently. ISO 9001 specifies requirements for a quality management system that can be used for internal application by organizations, or for certification, or for contractual purposes. It focuses on the effectiveness of the quality management system in meeting customer requirements. At the time of publication of this International Standard, ISO 9004 is under revision. The revised edition of ISO 9004 will provide guidance to management for achieving sustained success for any organization in a complex, demanding, and ever changing, environment. ISO 9004 provides a wider focus on quality management than ISO 9001; it addresses the needs and expectations of all interested parties and their satisfaction, by the systematic and continual improvement of the organization s performance. However, it is not intended for certification, regulatory or contractual use. 0.4 Compatibility with other management systems During the development of this International Standard, due consideration was given to the provisions of ISO 14001:2004 to enhance the compatibility of the two standards for the benefit of the user community. Annex A shows the correspondence between ISO 9001:2008 and ISO 14001:2004. This International Standard does not include requirements specific to other management systems, such as those particular to environmental management, occupational health and safety management, financial management or risk management. However, this International Standard enables an organization to align or integrate its own quality management system with related management system requirements. It is possible for an organization to adapt its existing management system(s) in order to establish a quality management system that complies with the requirements of this International Standard. ISO 2008 All rights reserved vii

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112 INTERNATIONAL STANDARD ISO 9001:2008(E) Quality management systems Requirements 1 Scope 1.1 General This International Standard specifies requirements for a quality management system where an organization a) needs to demonstrate its ability to consistently provide product that meets customer and applicable statutory and regulatory requirements, and b) aims to enhance customer satisfaction through the effective application of the system, including processes for continual improvement of the system and the assurance of conformity to customer and applicable statutory and regulatory requirements. NOTE 1 In this International Standard, the term product only applies to a) product intended for, or required by, a customer, b) any intended output resulting from the product realization processes. NOTE 2 Statutory and regulatory requirements can be expressed as legal requirements. 1.2 Application All requirements of this International Standard are generic and are intended to be applicable to all organizations, regardless of type, size and product provided. Where any requirement(s) of this International Standard cannot be applied due to the nature of an organization and its product, this can be considered for exclusion. Where exclusions are made, claims of conformity to this International Standard are not acceptable unless these exclusions are limited to requirements within Clause 7, and such exclusions do not affect the organization's ability, or responsibility, to provide product that meets customer and applicable statutory and regulatory requirements. 2 Normative references The following referenced documents are indispensable for the application of this document. For dated references, only the edition cited applies. For undated references, the latest edition of the referenced document (including any amendments) applies. ISO 9000:2005, Quality management systems Fundamentals and vocabulary 3 Terms and definitions For the purposes of this document, the terms and definitions given in ISO 9000 apply. Throughout the text of this International Standard, wherever the term product occurs, it can also mean service. ISO 2008 All rights reserved 1

113 ISO 9001:2008(E) 4 Quality management system 4.1 General requirements The organization shall establish, document, implement and maintain a quality management system and continually improve its effectiveness in accordance with the requirements of this International Standard. The organization shall a) determine the processes needed for the quality management system and their application throughout the organization (see 1.2), b) determine the sequence and interaction of these processes, c) determine criteria and methods needed to ensure that both the operation and control of these processes are effective, d) ensure the availability of resources and information necessary to support the operation and monitoring of these processes, e) monitor, measure where applicable, and analyse these processes, and f) implement actions necessary to achieve planned results and continual improvement of these processes. These processes shall be managed by the organization in accordance with the requirements of this International Standard. Where an organization chooses to outsource any process that affects product conformity to requirements, the organization shall ensure control over such processes. The type and extent of control to be applied to these outsourced processes shall be defined within the quality management system. NOTE 1 Processes needed for the quality management system referred to above include processes for management activities, provision of resources, product realization, measurement, analysis and improvement. NOTE 2 An outsourced process is a process that the organization needs for its quality management system and which the organization chooses to have performed by an external party. NOTE 3 Ensuring control over outsourced processes does not absolve the organization of the responsibility of conformity to all customer, statutory and regulatory requirements. The type and extent of control to be applied to the outsourced process can be influenced by factors such as a) the potential impact of the outsourced process on the organization's capability to provide product that conforms to requirements, b) the degree to which the control for the process is shared, c) the capability of achieving the necessary control through the application of Documentation requirements General The quality management system documentation shall include a) documented statements of a quality policy and quality objectives, b) a quality manual, c) documented procedures and records required by this International Standard, and d) documents, including records, determined by the organization to be necessary to ensure the effective planning, operation and control of its processes. 2 ISO 2008 All rights reserved

114 ISO 9001:2008(E) NOTE 1 Where the term documented procedure appears within this International Standard, this means that the procedure is established, documented, implemented and maintained. A single document may address the requirements for one or more procedures. A requirement for a documented procedure may be covered by more than one document. NOTE 2 The extent of the quality management system documentation can differ from one organization to another due to a) the size of organization and type of activities, b) the complexity of processes and their interactions, and c) the competence of personnel. NOTE 3 The documentation can be in any form or type of medium Quality manual The organization shall establish and maintain a quality manual that includes a) the scope of the quality management system, including details of and justification for any exclusions (see 1.2), b) the documented procedures established for the quality management system, or reference to them, and c) a description of the interaction between the processes of the quality management system Control of documents Documents required by the quality management system shall be controlled. Records are a special type of document and shall be controlled according to the requirements given in A documented procedure shall be established to define the controls needed a) to approve documents for adequacy prior to issue, b) to review and update as necessary and re-approve documents, c) to ensure that changes and the current revision status of documents are identified, d) to ensure that relevant versions of applicable documents are available at points of use, e) to ensure that documents remain legible and readily identifiable, f) to ensure that documents of external origin determined by the organization to be necessary for the planning and operation of the quality management system are identified and their distribution controlled, and g) to prevent the unintended use of obsolete documents, and to apply suitable identification to them if they are retained for any purpose Control of records Records established to provide evidence of conformity to requirements and of the effective operation of the quality management system shall be controlled. The organization shall establish a documented procedure to define the controls needed for the identification, storage, protection, retrieval, retention and disposition of records. Records shall remain legible, readily identifiable and retrievable. 5 Management responsibility 5.1 Management commitment Top management shall provide evidence of its commitment to the development and implementation of the quality management system and continually improving its effectiveness by ISO 2008 All rights reserved 3

115 ISO 9001:2008(E) a) communicating to the organization the importance of meeting customer as well as statutory and regulatory requirements, b) establishing the quality policy, c) ensuring that quality objectives are established, d) conducting management reviews, and e) ensuring the availability of resources. 5.2 Customer focus Top management shall ensure that customer requirements are determined and are met with the aim of enhancing customer satisfaction (see and 8.2.1). 5.3 Quality policy Top management shall ensure that the quality policy a) is appropriate to the purpose of the organization, b) includes a commitment to comply with requirements and continually improve the effectiveness of the quality management system, c) provides a framework for establishing and reviewing quality objectives, d) is communicated and understood within the organization, and e) is reviewed for continuing suitability. 5.4 Planning Quality objectives Top management shall ensure that quality objectives, including those needed to meet requirements for product [see 7.1 a)], are established at relevant functions and levels within the organization. The quality objectives shall be measurable and consistent with the quality policy Quality management system planning Top management shall ensure that a) the planning of the quality management system is carried out in order to meet the requirements given in 4.1, as well as the quality objectives, and b) the integrity of the quality management system is maintained when changes to the quality management system are planned and implemented. 5.5 Responsibility, authority and communication Responsibility and authority Top management shall ensure that responsibilities and authorities are defined and communicated within the organization. 4 ISO 2008 All rights reserved

116 ISO 9001:2008(E) Management representative Top management shall appoint a member of the organization's management who, irrespective of other responsibilities, shall have responsibility and authority that includes a) ensuring that processes needed for the quality management system are established, implemented and maintained, b) reporting to top management on the performance of the quality management system and any need for improvement, and c) ensuring the promotion of awareness of customer requirements throughout the organization. NOTE The responsibility of a management representative can include liaison with external parties on matters relating to the quality management system Internal communication Top management shall ensure that appropriate communication processes are established within the organization and that communication takes place regarding the effectiveness of the quality management system. 5.6 Management review General Top management shall review the organization's quality management system, at planned intervals, to ensure its continuing suitability, adequacy and effectiveness. This review shall include assessing opportunities for improvement and the need for changes to the quality management system, including the quality policy and quality objectives. Records from management reviews shall be maintained (see 4.2.4) Review input The input to management review shall include information on a) results of audits, b) customer feedback, c) process performance and product conformity, d) status of preventive and corrective actions, e) follow-up actions from previous management reviews, f) changes that could affect the quality management system, and g) recommendations for improvement Review output The output from the management review shall include any decisions and actions related to a) improvement of the effectiveness of the quality management system and its processes, b) improvement of product related to customer requirements, and c) resource needs. ISO 2008 All rights reserved 5

117 ISO 9001:2008(E) 6 Resource management 6.1 Provision of resources The organization shall determine and provide the resources needed a) to implement and maintain the quality management system and continually improve its effectiveness, and b) to enhance customer satisfaction by meeting customer requirements. 6.2 Human resources General Personnel performing work affecting conformity to product requirements shall be competent on the basis of appropriate education, training, skills and experience. NOTE Conformity to product requirements can be affected directly or indirectly by personnel performing any task within the quality management system Competence, training and awareness The organization shall a) determine the necessary competence for personnel performing work affecting conformity to product requirements, b) where applicable, provide training or take other actions to achieve the necessary competence, c) evaluate the effectiveness of the actions taken, d) ensure that its personnel are aware of the relevance and importance of their activities and how they contribute to the achievement of the quality objectives, and e) maintain appropriate records of education, training, skills and experience (see 4.2.4). 6.3 Infrastructure The organization shall determine, provide and maintain the infrastructure needed to achieve conformity to product requirements. Infrastructure includes, as applicable, a) buildings, workspace and associated utilities, b) process equipment (both hardware and software), and c) supporting services (such as transport, communication or information systems). 6.4 Work environment The organization shall determine and manage the work environment needed to achieve conformity to product requirements. NOTE The term work environment relates to those conditions under which work is performed including physical, environmental and other factors (such as noise, temperature, humidity, lighting or weather). 6 ISO 2008 All rights reserved

118 ISO 9001:2008(E) 7 Product realization 7.1 Planning of product realization The organization shall plan and develop the processes needed for product realization. Planning of product realization shall be consistent with the requirements of the other processes of the quality management system (see 4.1). In planning product realization, the organization shall determine the following, as appropriate: a) quality objectives and requirements for the product; b) the need to establish processes and documents, and to provide resources specific to the product; c) required verification, validation, monitoring, measurement, inspection and test activities specific to the product and the criteria for product acceptance; d) records needed to provide evidence that the realization processes and resulting product meet requirements (see 4.2.4). The output of this planning shall be in a form suitable for the organization's method of operations. NOTE 1 A document specifying the processes of the quality management system (including the product realization processes) and the resources to be applied to a specific product, project or contract can be referred to as a quality plan. NOTE 2 The organization may also apply the requirements given in 7.3 to the development of product realization processes. 7.2 Customer-related processes Determination of requirements related to the product The organization shall determine a) requirements specified by the customer, including the requirements for delivery and post-delivery activities, b) requirements not stated by the customer but necessary for specified or intended use, where known, c) statutory and regulatory requirements applicable to the product, and d) any additional requirements considered necessary by the organization. NOTE Post-delivery activities include, for example, actions under warranty provisions, contractual obligations such as maintenance services, and supplementary services such as recycling or final disposal Review of requirements related to the product The organization shall review the requirements related to the product. This review shall be conducted prior to the organization's commitment to supply a product to the customer (e.g. submission of tenders, acceptance of contracts or orders, acceptance of changes to contracts or orders) and shall ensure that a) product requirements are defined, b) contract or order requirements differing from those previously expressed are resolved, and c) the organization has the ability to meet the defined requirements. Records of the results of the review and actions arising from the review shall be maintained (see 4.2.4). Where the customer provides no documented statement of requirement, the customer requirements shall be confirmed by the organization before acceptance. ISO 2008 All rights reserved 7

119 ISO 9001:2008(E) Where product requirements are changed, the organization shall ensure that relevant documents are amended and that relevant personnel are made aware of the changed requirements. NOTE In some situations, such as internet sales, a formal review is impractical for each order. Instead the review can cover relevant product information such as catalogues or advertising material Customer communication The organization shall determine and implement effective arrangements for communicating with customers in relation to a) product information, b) enquiries, contracts or order handling, including amendments, and c) customer feedback, including customer complaints. 7.3 Design and development Design and development planning The organization shall plan and control the design and development of product. During the design and development planning, the organization shall determine a) the design and development stages, b) the review, verification and validation that are appropriate to each design and development stage, and c) the responsibilities and authorities for design and development. The organization shall manage the interfaces between different groups involved in design and development to ensure effective communication and clear assignment of responsibility. Planning output shall be updated, as appropriate, as the design and development progresses. NOTE Design and development review, verification and validation have distinct purposes. They can be conducted and recorded separately or in any combination, as suitable for the product and the organization Design and development inputs Inputs relating to product requirements shall be determined and records maintained (see 4.2.4). These inputs shall include a) functional and performance requirements, b) applicable statutory and regulatory requirements, c) where applicable, information derived from previous similar designs, and d) other requirements essential for design and development. The inputs shall be reviewed for adequacy. Requirements shall be complete, unambiguous and not in conflict with each other Design and development outputs The outputs of design and development shall be in a form suitable for verification against the design and development input and shall be approved prior to release. 8 ISO 2008 All rights reserved

120 ISO 9001:2008(E) Design and development outputs shall a) meet the input requirements for design and development, b) provide appropriate information for purchasing, production and service provision, c) contain or reference product acceptance criteria, and d) specify the characteristics of the product that are essential for its safe and proper use. NOTE Information for production and service provision can include details for the preservation of product Design and development review At suitable stages, systematic reviews of design and development shall be performed in accordance with planned arrangements (see 7.3.1) a) to evaluate the ability of the results of design and development to meet requirements, and b) to identify any problems and propose necessary actions. Participants in such reviews shall include representatives of functions concerned with the design and development stage(s) being reviewed. Records of the results of the reviews and any necessary actions shall be maintained (see 4.2.4) Design and development verification Verification shall be performed in accordance with planned arrangements (see 7.3.1) to ensure that the design and development outputs have met the design and development input requirements. Records of the results of the verification and any necessary actions shall be maintained (see 4.2.4) Design and development validation Design and development validation shall be performed in accordance with planned arrangements (see 7.3.1) to ensure that the resulting product is capable of meeting the requirements for the specified application or intended use, where known. Wherever practicable, validation shall be completed prior to the delivery or implementation of the product. Records of the results of validation and any necessary actions shall be maintained (see 4.2.4) Control of design and development changes Design and development changes shall be identified and records maintained. The changes shall be reviewed, verified and validated, as appropriate, and approved before implementation. The review of design and development changes shall include evaluation of the effect of the changes on constituent parts and product already delivered. Records of the results of the review of changes and any necessary actions shall be maintained (see 4.2.4). 7.4 Purchasing Purchasing process The organization shall ensure that purchased product conforms to specified purchase requirements. The type and extent of control applied to the supplier and the purchased product shall be dependent upon the effect of the purchased product on subsequent product realization or the final product. ISO 2008 All rights reserved 9

121 ISO 9001:2008(E) The organization shall evaluate and select suppliers based on their ability to supply product in accordance with the organization's requirements. Criteria for selection, evaluation and re-evaluation shall be established. Records of the results of evaluations and any necessary actions arising from the evaluation shall be maintained (see 4.2.4) Purchasing information Purchasing information shall describe the product to be purchased, including, where appropriate, a) requirements for approval of product, procedures, processes and equipment, b) requirements for qualification of personnel, and c) quality management system requirements. The organization shall ensure the adequacy of specified purchase requirements prior to their communication to the supplier Verification of purchased product The organization shall establish and implement the inspection or other activities necessary for ensuring that purchased product meets specified purchase requirements. Where the organization or its customer intends to perform verification at the supplier's premises, the organization shall state the intended verification arrangements and method of product release in the purchasing information. 7.5 Production and service provision Control of production and service provision The organization shall plan and carry out production and service provision under controlled conditions. Controlled conditions shall include, as applicable, a) the availability of information that describes the characteristics of the product, b) the availability of work instructions, as necessary, c) the use of suitable equipment, d) the availability and use of monitoring and measuring equipment, e) the implementation of monitoring and measurement, and f) the implementation of product release, delivery and post-delivery activities Validation of processes for production and service provision The organization shall validate any processes for production and service provision where the resulting output cannot be verified by subsequent monitoring or measurement and, as a consequence, deficiencies become apparent only after the product is in use or the service has been delivered. Validation shall demonstrate the ability of these processes to achieve planned results. The organization shall establish arrangements for these processes including, as applicable, a) defined criteria for review and approval of the processes, b) approval of equipment and qualification of personnel, c) use of specific methods and procedures, 10 ISO 2008 All rights reserved

122 ISO 9001:2008(E) d) requirements for records (see 4.2.4), and e) revalidation Identification and traceability Where appropriate, the organization shall identify the product by suitable means throughout product realization. The organization shall identify the product status with respect to monitoring and measurement requirements throughout product realization. Where traceability is a requirement, the organization shall control the unique identification of the product and maintain records (see 4.2.4). NOTE In some industry sectors, configuration management is a means by which identification and traceability are maintained Customer property The organization shall exercise care with customer property while it is under the organization's control or being used by the organization. The organization shall identify, verify, protect and safeguard customer property provided for use or incorporation into the product. If any customer property is lost, damaged or otherwise found to be unsuitable for use, the organization shall report this to the customer and maintain records (see 4.2.4). NOTE Customer property can include intellectual property and personal data Preservation of product The organization shall preserve the product during internal processing and delivery to the intended destination in order to maintain conformity to requirements. As applicable, preservation shall include identification, handling, packaging, storage and protection. Preservation shall also apply to the constituent parts of a product. 7.6 Control of monitoring and measuring equipment The organization shall determine the monitoring and measurement to be undertaken and the monitoring and measuring equipment needed to provide evidence of conformity of product to determined requirements. The organization shall establish processes to ensure that monitoring and measurement can be carried out and are carried out in a manner that is consistent with the monitoring and measurement requirements. Where necessary to ensure valid results, measuring equipment shall a) be calibrated or verified, or both, at specified intervals, or prior to use, against measurement standards traceable to international or national measurement standards; where no such standards exist, the basis used for calibration or verification shall be recorded (see 4.2.4); b) be adjusted or re-adjusted as necessary; c) have identification in order to determine its calibration status; d) be safeguarded from adjustments that would invalidate the measurement result; e) be protected from damage and deterioration during handling, maintenance and storage. In addition, the organization shall assess and record the validity of the previous measuring results when the equipment is found not to conform to requirements. The organization shall take appropriate action on the equipment and any product affected. Records of the results of calibration and verification shall be maintained (see 4.2.4). ISO 2008 All rights reserved 11

123 ISO 9001:2008(E) When used in the monitoring and measurement of specified requirements, the ability of computer software to satisfy the intended application shall be confirmed. This shall be undertaken prior to initial use and reconfirmed as necessary. NOTE Confirmation of the ability of computer software to satisfy the intended application would typically include its verification and configuration management to maintain its suitability for use. 8 Measurement, analysis and improvement 8.1 General The organization shall plan and implement the monitoring, measurement, analysis and improvement processes needed a) to demonstrate conformity to product requirements, b) to ensure conformity of the quality management system, and c) to continually improve the effectiveness of the quality management system. This shall include determination of applicable methods, including statistical techniques, and the extent of their use. 8.2 Monitoring and measurement Customer satisfaction As one of the measurements of the performance of the quality management system, the organization shall monitor information relating to customer perception as to whether the organization has met customer requirements. The methods for obtaining and using this information shall be determined. NOTE Monitoring customer perception can include obtaining input from sources such as customer satisfaction surveys, customer data on delivered product quality, user opinion surveys, lost business analysis, compliments, warranty claims and dealer reports Internal audit The organization shall conduct internal audits at planned intervals to determine whether the quality management system a) conforms to the planned arrangements (see 7.1), to the requirements of this International Standard and to the quality management system requirements established by the organization, and b) is effectively implemented and maintained. An audit programme shall be planned, taking into consideration the status and importance of the processes and areas to be audited, as well as the results of previous audits. The audit criteria, scope, frequency and methods shall be defined. The selection of auditors and conduct of audits shall ensure objectivity and impartiality of the audit process. Auditors shall not audit their own work. A documented procedure shall be established to define the responsibilities and requirements for planning and conducting audits, establishing records and reporting results. Records of the audits and their results shall be maintained (see 4.2.4). The management responsible for the area being audited shall ensure that any necessary corrections and corrective actions are taken without undue delay to eliminate detected nonconformities and their causes. 12 ISO 2008 All rights reserved

124 ISO 9001:2008(E) Follow-up activities shall include the verification of the actions taken and the reporting of verification results (see 8.5.2). NOTE See ISO for guidance Monitoring and measurement of processes The organization shall apply suitable methods for monitoring and, where applicable, measurement of the quality management system processes. These methods shall demonstrate the ability of the processes to achieve planned results. When planned results are not achieved, correction and corrective action shall be taken, as appropriate. NOTE When determining suitable methods, it is advisable that the organization consider the type and extent of monitoring or measurement appropriate to each of its processes in relation to their impact on the conformity to product requirements and on the effectiveness of the quality management system Monitoring and measurement of product The organization shall monitor and measure the characteristics of the product to verify that product requirements have been met. This shall be carried out at appropriate stages of the product realization process in accordance with the planned arrangements (see 7.1). Evidence of conformity with the acceptance criteria shall be maintained. Records shall indicate the person(s) authorizing release of product for delivery to the customer (see 4.2.4). The release of product and delivery of service to the customer shall not proceed until the planned arrangements (see 7.1) have been satisfactorily completed, unless otherwise approved by a relevant authority and, where applicable, by the customer. 8.3 Control of nonconforming product The organization shall ensure that product which does not conform to product requirements is identified and controlled to prevent its unintended use or delivery. A documented procedure shall be established to define the controls and related responsibilities and authorities for dealing with nonconforming product. Where applicable, the organization shall deal with nonconforming product by one or more of the following ways: a) by taking action to eliminate the detected nonconformity; b) by authorizing its use, release or acceptance under concession by a relevant authority and, where applicable, by the customer; c) by taking action to preclude its original intended use or application; d) by taking action appropriate to the effects, or potential effects, of the nonconformity when nonconforming product is detected after delivery or use has started. When nonconforming product is corrected it shall be subject to re-verification to demonstrate conformity to the requirements. Records of the nature of nonconformities and any subsequent actions taken, including concessions obtained, shall be maintained (see 4.2.4). 8.4 Analysis of data The organization shall determine, collect and analyse appropriate data to demonstrate the suitability and effectiveness of the quality management system and to evaluate where continual improvement of the effectiveness of the quality management system can be made. This shall include data generated as a result of monitoring and measurement and from other relevant sources. ISO 2008 All rights reserved 13

125 ISO 9001:2008(E) The analysis of data shall provide information relating to a) customer satisfaction (see 8.2.1), b) conformity to product requirements (see 8.2.4), c) characteristics and trends of processes and products, including opportunities for preventive action (see and 8.2.4), and d) suppliers (see 7.4). 8.5 Improvement Continual improvement The organization shall continually improve the effectiveness of the quality management system through the use of the quality policy, quality objectives, audit results, analysis of data, corrective and preventive actions and management review Corrective action The organization shall take action to eliminate the causes of nonconformities in order to prevent recurrence. Corrective actions shall be appropriate to the effects of the nonconformities encountered. A documented procedure shall be established to define requirements for a) reviewing nonconformities (including customer complaints), b) determining the causes of nonconformities, c) evaluating the need for action to ensure that nonconformities do not recur, d) determining and implementing action needed, e) records of the results of action taken (see 4.2.4), and f) reviewing the effectiveness of the corrective action taken Preventive action The organization shall determine action to eliminate the causes of potential nonconformities in order to prevent their occurrence. Preventive actions shall be appropriate to the effects of the potential problems. A documented procedure shall be established to define requirements for a) determining potential nonconformities and their causes, b) evaluating the need for action to prevent occurrence of nonconformities, c) determining and implementing action needed, d) records of results of action taken (see 4.2.4), and e) reviewing the effectiveness of the preventive action taken. 14 ISO 2008 All rights reserved

126 ISO 9001:2008(E) Annex A (informative) Correspondence between ISO 9001:2008 and ISO 14001:2004 Table A.1 Correspondence between ISO 9001:2008 and ISO 14001:2004 ISO 9001:2008 ISO 14001:2004 Introduction (title only) Introduction General 0.1 Process approach 0.2 Relationship with ISO Compatibility with other management systems 0.4 Scope (title only) 1 1 Scope General 1.1 Application 1.2 Normative references 2 2 Normative references Terms and definitions 3 3 Terms and definitions Quality management system (title only) 4 4 Environmental management system requirements (title only) General requirements General requirements Documentation requirements (title only) 4.2 General Documentation Quality manual Control of documents Control of documents Control of records Control of records Management responsibility (title only) 5 Management commitment Environmental policy Resources, roles, responsibility and authority Customer focus Environmental aspects Legal and other requirements 4.6 Management review Quality policy Environmental policy Planning (title only) Planning (title only) Quality objectives Objectives, targets and programme(s) Quality management system planning Objectives, targets and programme(s) Responsibility, authority and communication (title 5.5 only) Responsibility and authority General requirements Resources, roles, responsibility and authority Management representative Resources, roles, responsibility and authority Internal communication Communication Management review (title only) Management review General Management review Review input Management review Review output Management review ISO 2008 All rights reserved 15

127 ISO 9001:2008(E) Table A.1 Correspondence between ISO 9001:2008 and ISO 14001:2004 (continued) ISO 9001:2008 ISO 14001:2004 Resource management (title only) 6 Provision of resources Resources, roles, responsibility and authority Human resources (title only) 6.2 General Competence, training and awareness Competence, training and awareness Competence, training and awareness Infrastructure Resources, roles, responsibility and authority Work environment 6.4 Product realization (title only) Implementation and operation (title only) Planning of product realization Operational control Customer-related processes (title only) 7.2 Determination of requirements related to the Environmental aspects product Legal and other requirements Operational control Review of requirements related to the product Environmental aspects Operational control Customer communication Communication Design and development (title only) 7.3 Design and development planning Operational control Design and development inputs Operational control Design and development outputs Operational control Design and development review Operational control Design and development verification Operational control Design and development validation Operational control Control of design and development changes Operational control Purchasing (title only) 7.4 Purchasing process Operational control Purchasing information Operational control Verification of purchased product Operational control Production and service provision (title only) 7.5 Control of production and service provision Operational control Validation of processes for production and service Operational control provision Identification and traceability Customer property Preservation of product Operational control Control of monitoring and measuring equipment Monitoring and measurement Measurement, analysis and improvement (title only) Checking (title only) General Monitoring and measurement Monitoring and measurement (title only) 8.2 Customer satisfaction Internal audit Internal audit Monitoring and measurement of processes Monitoring and measurement Evaluation of compliance 16 ISO 2008 All rights reserved

128 ISO 9001:2008(E) Table A.1 Correspondence between ISO 9001:2008 and ISO 14001:2004 (continued) ISO 9001:2008 ISO 14001:2004 Monitoring and measurement of product Monitoring and measurement Evaluation of compliance Control of nonconforming product Emergency preparedness and response Nonconformity, corrective action and preventive action Analysis of data Monitoring and measurement Improvement (title only) 8.5 Continual improvement Environmental policy Objectives, targets and programme(s) 4.6 Management review Corrective action Nonconformity, corrective action and preventive action Preventive action Nonconformity, corrective action and preventive action ISO 2008 All rights reserved 17

129 ISO 9001:2008(E) Table A.2 Correspondence between ISO 14001:2004 and ISO 9001:2008 ISO 14001:2004 ISO 9001:2008 Introduction Introduction (title only) 0.1 General 0.2 Process approach 0.3 Relationship with ISO Compatibility with other management systems Scope 1 1 Scope (title only) 1.1 General 1.2 Application Normative references 2 2 Normative references Terms and definitions 3 3 Terms and definitions Environmental management system requirements 4 4 Quality management system (title only) (title only) General requirements General requirements 5.5 Responsibility, authority and communication (title only) Responsibility and authority Environmental policy Management commitment 5.3 Quality policy Continual improvement Planning (title only) Planning (title only) Environmental aspects Customer focus Determination of requirements related to the product Review of requirements related to the product Legal and other requirements Customer focus Determination of requirements related to the product Objectives, targets and programme(s) Quality objectives Quality management system planning Continual improvement Implementation and operation (title only) Product realization (title only) Resources, roles, responsibility and authority Management commitment Responsibility and authority Management representative 6.1 Provision of resources 6.3 Infrastructure Competence, training and awareness (Human resources) General Competence, training and awareness Communication Internal communication Customer communication Documentation (Documentation requirements) General Control of documents Control of documents 18 ISO 2008 All rights reserved

130 ISO 9001:2008(E) Table A.2 Correspondence between ISO 14001:2004 and ISO 9001:2008 (continued) ISO 14001:2004 ISO 9001:2008 Operational control Planning of product realization 7.2 Customer-related processes (title only) Determination of requirements related to the product Review of requirements related to the product Design and development planning Design and development inputs Design and development outputs Design and development review Design and development verification Design and development validation Control of design and development changes Purchasing process Purchasing information Verification of purchased product 7.5 Production and service provision (title only) Control of production and service provision Validation of processes for production and service provision Preservation of product Emergency preparedness and response Control of nonconforming product Checking (title only) Measurement, analysis and improvement (title only) Monitoring and measurement Control of monitoring and measuring equipment 8.1 (Measurement, analysis and improvement) General Monitoring and measurement of processes Monitoring and measurement of product 8.4 Analysis of data Evaluation of compliance Monitoring and measurement of processes Monitoring and measurement of product Nonconformity, corrective action and preventive Control of nonconforming product action 8.4 Analysis of data Corrective action Preventive action Control of records Control of records Internal audit Internal audit Management review Management commitment 5.6 Management review (title only) General Review input Review output Continual improvement ISO 2008 All rights reserved 19

131 ISO 9001:2008(E) Annex B (informative) Changes between ISO 9001:2000 and ISO 9001:2008 Table B.1 Changes between ISO 9001:2000 and ISO 9001:2008 ISO 9001:2000 Clause No. Paragraph/ Figure/ Table/ Note Addition (A) or Deletion (D) Amended text Foreword Para 2 D + A International Standards are drafted in accordance with the rules given in the ISO/IEC Directives, Part 3 Part 2. Foreword Para 3, Sentence 1 Foreword Para 4, Sentence 1 A D + A The main task of technical committees is to prepare International Standards. Attention is drawn to the possibility that some of the elements of this International Standard document may be the subject of patent rights. Foreword Para 5 D International Standard ISO 9001 was prepared by Technical Committee ISO/TC 176, Quality management and quality assurance, Subcommittee SC 2, Quality systems. Foreword Para 6 D This third edition of ISO 9001 cancels and replaces the second edition (ISO 9001:1994) together with ISO 9002:1994 and ISO 9003:1994. It constitutes a technical revision of these documents. Those organizations which have used ISO 9002:1994 and ISO 9003:1994 in the past may use this International Standard by excluding certain requirements in accordance with 1.2. A This fourth edition cancels and replaces the third edition (ISO 9001:2000), which has been amended to clarify points in the text and to enhance compatibility with ISO 14001:2004. Foreword Para 7 D The title of ISO 9001 has been revised in this edition and no longer includes the term Quality assurance. This reflects the fact that the quality management system requirements specified in this edition of ISO 9001, in addition to quality assurance of product, also aim to enhance customer satisfaction. Foreword Para 8 D Annexes A and B of this International Standard are for information only. Foreword New para 7 A Details of the changes between the third edition and this fourth edition are given in Annex B. 0.1 Para 1, Sentence 2 D A The design and implementation of an organization's quality management system is influenced by varying needs, particular objectives, the products provided, the processes employed and the size and structure of the organization. The design and implementation of an organization's quality management system is influenced by a) its organizational environment, change in that environment, and the risks associated with that environment; b) its varying needs; c) its particular objectives; d) the products it provides; e) the processes it employs; f) its size and organizational structure. Sentence 3 Now a new para It is not the intent of this International Standard to imply uniformity in the structure of quality management systems or uniformity of documentation. 0.1 Para 4 A This International Standard can be used by internal and external parties, including certification bodies, to assess the organization's ability to meet customer, statutory and regulatory requirements applicable to the product, and the organization's own requirements. 0.2 Para 2 D + A For an organization to function effectively, it has to identify determine and manage numerous linked activities. An activity or set of activities using resources, and managed in order to enable the transformation of inputs into outputs, can be considered as a process. 20 ISO 2008 All rights reserved

132 ISO 9001:2008(E) Table B.1 Changes between ISO 9001:2000 and ISO 9001:2008 (continued) ISO 9001:2000 Clause No. Paragraph/ Figure/ Table/ Note Addition (A) or Deletion (D) Amended text 0.2 Para 3 A The application of a system of processes within an organization, together with the identification and interactions of these processes, and their management to produce the desired outcome, can be referred to as the process approach. 0.3 Para 1 D + A The present editions of ISO 9001 and ISO 9004 have been developed as a consistent pair of are quality management system standards which have been designed to complement each other, but can also be used independently. Although the two International Standards have different scopes, they have similar structures in order to assist their application as a consistent pair. 0.3 Para 3 D + A ISO 9004 gives a guidance on a wider range of objectives of a quality management system than does ISO 9001, particularly for the continual improvement of an organization s overall performance and efficiency, as well as its effectiveness. ISO 9004 is recommended as a guide for organizations whose top management wishes to move beyond the requirements of ISO 9001, in pursuit of continual improvement of performance. However, it is not intended for certification or for contractual purposes. At the time of publication of this International Standard, ISO 9004 is under revision. The revised edition of ISO 9004 will provide guidance to management for achieving sustained success for any organization in a complex, demanding, and ever changing, environment. ISO 9004 provides a wider focus on quality management than ISO 9001; it addresses the needs and expectations of all interested parties and their satisfaction, by the systematic and continual improvement of the organization s performance. However, it is not intended for certification, regulatory or contractual use. 0.4 Para 1 D + A This International Standard has been aligned with ISO 14001:1996 in order to enhance the compatibility of the two standards for the benefit of the user community. During the development of this International Standard, due consideration was given to the provisions of ISO 14001:2004 to enhance the compatibility of the two standards for the benefit of the user community. Annex A shows the correspondence between ISO 9001:2008 and ISO 14001: Bullet a) A a) needs to demonstrate its ability to consistently provide product that meets customer and applicable statutory and regulatory requirements, and Bullet b) A b) aims to enhance customer satisfaction through the effective application of the system, including processes for continual improvement of the system and the assurance of conformity to customer and applicable statutory and regulatory requirements. Note D NOTE In this International Standard, the term product applies only to the product intended for, or required by, a customer. A NOTE 1 In this International Standard, the term product only applies to a) a product intended for, or required by, a customer, b) any intended output resulting from the product realization processes. New Note 2 A NOTE 2 Statutory and regulatory requirements can be expressed as legal requirements. 1.2 Para 3 A Where exclusions are made, claims of conformity to this International Standard are not acceptable unless these exclusions are limited to requirements within Clause 7, and such exclusions do not affect the organization's ability, or responsibility, to provide product that meets customer and applicable statutory and regulatory requirements. 2 Para 1 D + A The following normative document contains provisions which, through reference in this text, constitute provisions of this International Standard. For dated references, subsequent amendments to, or revisions of, any of these publications do not apply. However, parties to agreements based on this International Standard are encouraged to investigate the possibility of applying the most recent edition of the normative document indicated below. For undated references, the latest edition of the normative document referred to applies. Members of ISO and IEC maintain registers of currently valid International Standards. A D + A The following referenced documents are indispensable for the application of this document. For dated references, only the edition cited applies. For undated references, the latest edition of the referenced document (including any amendments) applies. ISO 9000: , Quality management systems Fundamentals and vocabulary ISO 2008 All rights reserved 21

133 ISO 9001:2008(E) Table B.1 Changes between ISO 9001:2000 and ISO 9001:2008 (continued) ISO 9001:2000 Clause No. Paragraph/ Figure/ Table/ Note Addition (A) or Deletion (D) Amended text 3 Para 1 D + A For the purposes of this document International Standard, the terms and definitions given in ISO 9000 apply. 3 Paras 2, 3 D The following terms, used in this edition of ISO 9001 to describe the supply chain, have been changed to reflect the vocabulary currently used: supplier organization customer The term organization replaces the term supplier used in ISO 9001:1994, and refers to the unit to which this International Standard applies. Also, the term supplier now replaces the term subcontractor. 4.1 Bullet a) D + A a) identify determine the processes needed for the quality management system and their application throughout the organization (see 1.2), 4.1 Bullet e) A e) monitor, measure where applicable, and analyse these processes, and 4.1 Para 4 D + A Where an organization chooses to outsource any process that affects product conformity with to requirements, the organization shall ensure control over such processes. The type and extent of control to be applied to these outsourced processes shall be defined within the quality management system. 4.1 Note 1 D + A NOTE 1 Processes needed for the quality management system referred to above should include processes for management activities, provision of resources, product realization, and measurement, analysis and improvement. 4.1 New Notes 2 & 3 A NOTE 2 An outsourced process is a process that the organization needs for its quality management system and which the organization chooses to have performed by an external party. NOTE 3 Ensuring control over outsourced processes does not absolve the organization of the responsibility of conformity to all customer, statutory and regulatory requirements. The type and extent of control to be applied to the outsourced process can be influenced by factors such as a) the potential impact of the outsourced process on the organization's capability to provide product that conforms to requirements, b) the degree to which the control for the process is shared, c) the capability of achieving the necessary control through the application of Bullet c) A c) documented procedures and records required by this International Standard, and Bullet d) A + D d) documents, including records, needed determined by the organization to be necessary to ensure the effective planning, operation and control of its processes. and Bullet e) D e) records required by this International Standard (see 4.2.4) Note 1 A NOTE 1 Where the term documented procedure appears within this International Standard, this means that the procedure is established, documented, implemented and maintained. A single document may address the requirements for one or more procedures. A requirement for a documented procedure may be covered by more than one document Bullet f) A f) to ensure that documents of external origin determined by the organization to be necessary for the planning and operation of the quality management system are identified and their distribution controlled, and Para 1 D + A Records shall be established and maintained to provide evidence of conformity to requirements and of the effective operation of the quality management system shall be controlled. Records shall remain legible, readily identifiable and retrievable. The organization shall establish a documented procedure shall be established to define the controls needed for the identification, storage, protection, retrieval, retention time and disposition of records. Records shall remain legible, readily identifiable and retrievable Para 1 A Top management shall appoint a member of the organization's management who, irrespective of other responsibilities, shall have responsibility and authority that includes Para 1 A + D Personnel performing work affecting conformity to product quality requirements shall be competent on the basis of appropriate education, training, skills and experience. New Note A NOTE Conformity to product requirements can be affected directly or indirectly by personnel performing any task within the quality management system. 22 ISO 2008 All rights reserved

134 ISO 9001:2008(E) Table B.1 Changes between ISO 9001:2000 and ISO 9001:2008 (continued) ISO 9001:2000 Clause No. Paragraph/ Figure/ Table/ Note Addition (A) or Deletion (D) Amended text Clause title A + D Competence, training and awareness and training Bullets a) & b) A + D a) determine the necessary competence for personnel performing work affecting conformity to product quality requirements, b) where applicable, provide training or take other actions to satisfy these needs achieve the necessary competence, 6.3 Bullet c) A c) supporting services (such as transport, communication or information systems). 6.4 New Note A NOTE The term work environment relates to those conditions under which work is performed including physical, environmental and other factors (such as noise, temperature, humidity, lighting or weather). 7.1 Bullet b) A + D b) the need to establish processes and documents, and to provide resources specific to the product; 7.1 Bullet c) A c) required verification, validation, monitoring, measurement, inspection and test activities specific to the product and the criteria for product acceptance; Bullet c) Bullet d), New Note D+A D+A A c) statutory and regulatory requirements related applicable to the product, and d) any additional requirements determined considered necessary by the organization. NOTE Post-delivery activities include, for example, actions under warranty provisions, contractual obligations such as maintenance services, and supplementary services such as recycling or final disposal New Note A NOTE Design and development review, verification and validation have distinct purposes. They can be conducted and recorded separately or in any combination, as suitable for the product and the organization Para 2 D + A These The inputs shall be reviewed for adequacy. Requirements shall be complete, unambiguous and not in conflict with each other Para 1 D + A The outputs of design and development shall be provided in a form that enables in a form suitable for verification against the design and development input and shall be approved prior to release Bullet b) D b) provide appropriate information for purchasing, production and for service provision, New Note A NOTE Information for production and service provision can include details for the preservation of product Paras 1 & 2 No text change. Paras now merged Design and development changes shall be identified and records maintained. The changes shall be reviewed, verified and validated, as appropriate, and approved before implementation. The review of design and development changes shall include evaluation of the effect of the changes on constituent parts and product already delivered. Records of the results of the review of changes and any necessary actions shall be maintained (see 4.2.4) Bullet d) D + A d) the availability and use of monitoring and measuring devices equipment, Bullet f) A f) the implementation of product release, delivery and post-delivery activities Para 1 D + A The organization shall validate any processes for production and service provision where the resulting output cannot be verified by subsequent monitoring or measurement This includes any processes where and, as a consequence, deficiencies become apparent only after the product is in use or the service has been delivered Para 2 A The organization shall identify the product status with respect to monitoring and measurement requirements throughout product realization Para 3 D + A Where traceability is a requirement, the organization shall control and record the unique identification of the product and maintain records (see 4.2.4) Para 1, Sentence 3 D + A If any customer property is lost, damaged or otherwise found to be unsuitable for use, this shall be reported to the customer and records maintained the organization shall report this to the customer and maintain records (see 4.2.4). Note A NOTE Customer property can include intellectual property and personal data. ISO 2008 All rights reserved 23

135 ISO 9001:2008(E) Table B.1 Changes between ISO 9001:2000 and ISO 9001:2008 (continued) ISO 9001:2000 Clause No. Paragraph/ Figure/ Table/ Note Addition (A) or Deletion (D) Amended text Para 1 D + A The organization shall preserve the conformity of product during internal processing and delivery to the intended destination in order to maintain conformity to requirements. This As applicable, preservation shall include identification, handling, packaging, storage and protection. Preservation shall also apply to the constituent parts of a product. 7.6 Title D + A Control of monitoring and measuring devices equipment 7.6 Para 1 D + A The organization shall determine the monitoring and measurement to be undertaken and the monitoring and measuring devices equipment needed to provide evidence of conformity of product to determined requirements (see 7.2.1). 7.6 Bullet a) A a) be calibrated or verified, or both, at specified intervals, or prior to use, against measurement standards traceable to international or national measurement standards; where no such standards exist, the basis used for calibration or verification shall be recorded (see 4.2.4); 7.6 Bullet c) D + A c) be identified to enable the calibration status to be determined; c) have identification in order to determine its calibration status; 7.6 Para 4, Sentence 3 Now new para 5, without change. Records of the results of calibration and verification shall be maintained (see 4.2.4). 7.6 Note D + A NOTE See ISO and ISO for guidance NOTE Confirmation of the ability of computer software to satisfy the intended application would typically include its verification and configuration management to maintain its suitability for use. 8.1 Bullet a) D + A a) to demonstrate conformity of the product to product requirements, New Note A NOTE Monitoring customer perception can include obtaining input from sources such as customer satisfaction surveys, customer data on delivered product quality, user opinion surveys, lost business analysis, compliments, warranty claims and dealer reports Para 2 Sentence 3 A The selection of auditors and conduct of audits shall ensure objectivity and impartiality of the audit process New Para 3 A A documented procedure shall be established to define the responsibilities and requirements for planning and conducting audits, establishing records and reporting results Para 3 Now para 4 D+A Para 4 Sentence 1 Now para 5 A The responsibilities and requirements for planning and conducting audits, and for reporting results and maintaining records (see 4.2.4) shall be defined in a documented procedure. Records of the audits and their results shall be maintained (see 4.2.4). The management responsible for the area being audited shall ensure that any necessary corrections and corrective actions are taken without undue delay to eliminate detected nonconformities and their causes Note D + A NOTE See ISO , ISO and ISO See ISO for guidance Para 1 Sentence 3 D When planned results are not achieved, correction and corrective action shall be taken, as appropriate, to ensure conformity of the product New Note A NOTE When determining suitable methods, it is advisable that the organization consider the type and extent of monitoring or measurement appropriate to each of its processes in relation to their impact on the conformity to product requirements and on the effectiveness of the quality management system Para 1 A The organization shall monitor and measure the characteristics of the product to verify that product requirements have been met. This shall be carried out at appropriate stages of the product realization process in accordance with the planned arrangements (see 7.1). Evidence of conformity with the acceptance criteria shall be maintained. Para 2 D + A Evidence of conformity with the acceptance criteria shall be maintained. Records shall indicate the person(s) authorizing release of product for delivery to the customer (see 4.2.4). Para 3 D + A Product release and service delivery The release of product and delivery of service to the customer shall not proceed until the planned arrangements (see 7.1) have been satisfactorily completed, unless otherwise approved by a relevant authority and, where applicable, by the customer. 24 ISO 2008 All rights reserved

136 ISO 9001:2008(E) Table B.1 Changes between ISO 9001:2000 and ISO 9001:2008 (continued) ISO 9001:2000 Clause No. Paragraph/ Figure/ Table/ Note Addition (A) or Deletion (D) Amended text 8.3 Para 1, Sentence 2 D + A The controls and related responsibilities and authorities for dealing with nonconforming product shall be defined in a documented procedure. A documented procedure shall be established to define the controls and related responsibilities and authorities for dealing with nonconforming product. 8.3 Para 2 A Where applicable, the organization shall deal with nonconforming product by one or more of the following ways: 8.3 New bullet d) Para 3 Para 4 Para 5 A d) by taking action appropriate to the effects, or potential effects, of the nonconformity when nonconforming product is detected after delivery or use has started. Moved to Records of the nature of nonconformities and any subsequent actions taken, including be Para 4 concessions obtained, shall be maintained (see 4.2.4) Moved to When nonconforming product is corrected it shall be subject to re-verification to demonstrate be Para 3 conformity to the requirements. Records of the nature of nonconformities and any subsequent actions taken, including concessions obtained, shall be maintained (see 4.2.4). Now new bullet d) When nonconforming product is detected after delivery or use has started, the organization shall take action appropriate to the effects, or potential effects, of the nonconformity. 8.4 Bullet b) D + A b) conformity to product requirements (see 7.2.1) (see 8.2.4), Bullet c) A c) characteristics and trends of processes and products, including opportunities for preventive action (see and 8.2.4), and Bullet d) A d) suppliers (see 7.4) Para 1 D + A The organization shall take action to eliminate the cause causes of nonconformities in order to prevent recurrence Bullet f) A f) reviewing the effectiveness of the corrective action taken Bullet e) A e) reviewing the effectiveness of the preventive action taken. Annex A All D + A Updated to reflect ISO 9001:2008 versus ISO 14001:2004 Annex B All D + A Updated to reflect ISO 9001:2008 versus ISO 9001:2000 Bibliography New and amended references D+A Updated to reflect new standards (including ISO 9004, currently under revision), new editions of standards, or withdrawn standards. ISO 2008 All rights reserved 25

137 ISO 9001:2008(E) Bibliography [1] ISO 9004: 1), Managing for the sustained success of an organization A quality management approach [2] ISO 10001:2007, Quality management Customer satisfaction Guidelines for codes of conduct for organizations [3] ISO 10002:2004, Quality management Customer satisfaction Guidelines for complaints handling in organizations [4] ISO 10003:2007, Quality management Customer satisfaction Guidelines for dispute resolution external to organizations [5] ISO 10005:2005, Quality management systems Guidelines for quality plans [6] ISO 10006:2003, Quality management systems Guidelines for quality management in projects [7] ISO 10007:2003, Quality management systems Guidelines for configuration management [8] ISO 10012:2003, Measurement management systems Requirements for measurement processes and measuring equipment [9] ISO/TR 10013:2001, Guidelines for quality management system documentation [10] ISO 10014:2006, Quality management Guidelines for realizing financial and economic benefits [11] ISO 10015:1999, Quality management Guidelines for training [12] ISO/TR 10017:2003, Guidance on statistical techniques for ISO 9001:2000 [13] ISO 10019:2005, Guidelines for the selection of quality management system consultants and use of their services [14] ISO 14001:2004, Environmental management systems Requirements with guidance for use [15] ISO 19011:2002, Guidelines for quality and/or environmental management systems auditing [16] IEC :2003, Dependability management Part 1: Dependability management systems [17] IEC 61160:2006, Design review [18] ISO/IEC 90003:2004, Software engineering Guidelines for the application of ISO 9001:2000 to computer software [19] Quality management principles 2), ISO, 2001 [20] ISO 9000 Selection and use 2), ISO, 2008 [21] ISO 9001 for Small Businesses What to do; Advice from ISO/TC 176 3), ISO, ) To be published. (Revision of ISO 9004:2000) 2) Available from website: 3) To be updated and aligned with ISO 9001: ISO 2008 All rights reserved

138 ISO 9001:2008(E) [22] ISO Management Systems 4) [23] Reference web sites: ) A bimonthly publication which provides comprehensive coverage of international developments relating to ISO's management system standards, including news of their implementation by diverse organizations around the world. Available from ISO Central Secretariat ([email protected]). ISO 2008 All rights reserved 27

139 ISO 9001:2008(E) ICS Price based on 27 pages ISO 2008 All rights reserved

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147 BIO-ETS Q&A Q: What is BIO-ETS? A: BIO-ETS is the Excellence Through Stewardship (ETS) quality management program created by the Biotechnology Industry Organization (BIO). It is the first industry-coordinated effort to promote the responsible use of agricultural biotechnology, the global adoption of plant biotechnology and the enhanced value of biotech-derived plant products in the marketplace. Pioneer has made the commitment to adopt and adhere to the principles and best practices of BIO-ETS as a member company. Q: How does BIO-ETS work with existing Quality Management Systems (ISO 9000, GLP, etc.)? A: Any quality management program (QMP) we implement at Pioneer whether it be ISO 9000, BIO-ETS or anything else does not tell us what we need to do in our research activities, but instead offers guidelines or best management practices. We, as the research organization, describe our activities and then utilize the QMP to help organize and describe our process, along with conducting proper monitoring and documentation. With this in mind, we have decided to establish the Research Management System our overall QMP, which will utilize industry standards such as BIO-ETS and ISO 9000 to ensure our research processes operate effectively to meet business requirements (our processes are under control ). Q: What are the key points we need to consider regarding implementation of BIO-ETS? A: Once we describe our research processes, we need to review them to ensure we have addressed all the possible areas involved with Plant Product Integrity (PPI). PPI is the specific identity of a plant and purity of populations of that plant, which are established and maintained using appropriate measures. Another key is to describe the Critical Control Points (CCP), a step at which control can be applied and is essential to prevent, eliminate or reduce to an acceptable level, an activity that may compromise PPI. Q: What is in it for DuPont/Pioneer to implement BIO-ETS? A: The stated objectives of the BIO-ETS program are: 1. Full compliance by member companies with applicable regulatory requirements 2. Member company plant product integrity 3. The flow of goods in commerce and the prevention of trade disruptions. From an industry perspective the evidence of BIO-ETS implementation will promote the acceptance of biotechnology products around the world. Q: What impact will BIO-ETS have on our daily research activities? A: As discussed above we will establish our Research Management System which describes our process and activities. Our intent is to create a system that that is simple yet effective. It will have minimal administrative requirements while still doing an excellent job of describing our processes/activities, which will allow us to identify and implement continuous improvement opportunities.

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149 Page 1 of 1 RECIBO BANCARIO DE PAGO DE CONTRIBUCIONES, PRODUCTOS Y APROVECHAMIENTOS FEDERALES RFC: PHI931220AN5 Denominación o Razón Social: PHI MEXICO SA DE CV Fecha y Hora de Pago: 30/04/2009 4:28:00 P.M. Total de Pagos: 1 No. Autorización del medio: No. Operación: Página: 1/1 Llave de Pago: D601FAEE45 TOTAL EFECTIVAMENTE PAGADO: $ 3,636 Por los siguientes conceptos: 1/1 DERECHOS, PRODUCTOS Y APROVECHAMIENTOS Dependencia:11 Secretaría de Agricultura, Ganadería, Desarrollo Rural, Pesca y Alimentación. Clave de Referencia del DPA: Cadena de la dependencia: dgia002 Importe:3,636 Cantidad Pagada 3,636 Cadena 10001=PHI931220AN = = = = =16: =D601FAEE = = = = = DGIA = Sello Digital k9lyzoaxi4ma+dys3swxcqsxz/46oedmryzy/inqrmxj/dhp8opro2yl44qljsnldtsql2xi/zreyz8u6ciejldnamkhlaxfphmxvd/t8 Psl/CNQaHDR461LTckmqCQpc1WQ5RWJucWKP3itBku/kRhhozjHpyEbKuQrKYEVxOE= 30/04/2009