Chapter 9. Microbial Genetics. Copyright The McGraw-Hill Companies, Inc. Permission required for reproduction or display.

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1 Chapter 9 Microbial Genetics

2 Genetics and Genes Genetics the study of heredity The science of genetics explores: 1. Transmission of biological traits from parent to offspring 2. Expression and variation of those traits 3. Structure and function of genetic material 4. How this material changes 2

3 Levels of Structure and Function of the Genome Genome sum total of genetic material of a cell (chromosomes + mitochondria/chloroplasts and/or plasmids) Genome of cells DNA Genome of viruses DNA or RNA DNA complexed with protein constitutes the genetic material as chromosomes Organism level Cell level Chromosome level Molecular level T A G C A T C C G T A G C A T A G C C G PhotoLink/Photodisc/Getty Images RF A T CG A 3 T G C C G

4 Microbial Genomes Bacterial chromosomes are a single circular loop Eukaryotic chromosomes are multiple and linear Eukaryote (composite) Cells Chromosomes Prokaryote Nucleus Nucleolus Mitochondrion Chromosome Plasmids Plasmid (in some fungi and protozoa) Extrachromosomal DNA Viruses Chloroplast DNA RNA 4

5 Genes Chromosome is subdivided into genes, the fundamental unit of heredity responsible for a given trait Site on the chromosome that provides information for a certain cell function Segment of DNA that contains the necessary code to make a protein or RNA molecule Three basic categories of genes: 1. Genes that code for proteins structural genes 2. Genes that code for RNA 3. Genes that control gene expression regulatory genes 5

6 Genotypes and Phenotypes All types of genes constitute the genetic makeup genotype The expression of the genotype creates observable traits phenotype 6

7 Genomes Vary in Size Smallest virus 4-5 genes E. coli single chromosome containing 4,288 genes; 1 mm; 1,000X longer than cell Human cell 46 chromosomes containing 31,000 genes; 6 feet; 180,000X longer than cell 7

8 DNA Two strands twisted into a double helix Basic unit of DNA structure is a nucleotide Each nucleotide consists of 3 parts: A 5 carbon sugar deoxyribose A phosphate group A nitrogenous base adenine, guanine, thymine, cytosine Nucleotides covalently bond to form a sugar-phosphate backbone Each sugar attaches to two phosphates 5 carbon and 3 carbon 8

9 Deoxyribose sugar N base D Backbone D P D A C DNA Hydrogen bonds T G D D P P Phosphate P P D G C D P Condensed metaphase chromosome DNA DNA double helix DNA wrapped around histones with linker DNA between them DNA Histone Chemical tags attached to histoneproteins may increase the expression of nearby genes. Nucleosome Supercoiled condensed chromatin Condensed nucleosomes Loosely condensed chromosome Chromatin Uncondensed chromatin fiber 9

10 DNA Nitrogenous bases covalently bond to the 1 carbon of each sugar and span the center of the molecule to pair with an appropriate complementary base on the other strand Adenine binds to thymine with 2 hydrogen bonds Guanine binds to cytosine with 3 hydrogen bonds Antiparallel strands 3 to 5 and 5 to 3 H N Sugar Sugar phosphate G N-H N NH H-N Nitrogen base pair C N O Sugar phosphate P 3 5 OH 4 D P 5 P Phosphate P Deoxyribose 3 D C 2 with carbon number G C Cytosine H OH N D Sugar N N H C H O A N N H C H H N-H O CH 3 H-N T N O H P H G T A Guanine Thymin e Adenine Hydrogen bond Covalent bond 10 (a)

11 The Information in DNA Each strand provides a template for the exact copying of a new strand Order of bases constitutes the DNA code 11

12 H H N O H-N H N G N-H C N N Sugar NH O H H Sugar phosphate Nitrogen base pair Sugar phosphate P 5 4 D P C G 3 OH 5 P Phosphate P 5 4 Deoxyribose 3 D 1 2 with carbon number C Cytosine 5 3 Base pairs Sugar phosphate backbone C G Guanine T Thymin e Minor groove OH D C P A Adenine Hydrogen bond Covalent bond (b) 5 3 Major groove (c) (a) H H N N-H O CH 3 N A N H-N T H N N Sugar H O 12

13 Significance of DNA Structure 1. Maintenance of code during reproduction - Constancy of base pairing guarantees that the code will be retained 2. Providing variety - order of bases responsible for unique qualities of each organism 13

14 Concept Check: In DNA, adenine is the complementary base for, and cytosine is the complement for. A. guanine, thymine B. uracil, guanine C. thymine, guanine D. thymine, uracil

15 The Overall Replication Process Replication occurs on both strands simultaneously G C 5 3 G C A T T A T A G C A T Parental helix Creates complementary strands A T G C C C T A G C G A T G Replication fork G G Semiconservative replication process A T A T T A C G C T A A T A T A T T A C G C T A A T 3 G C 5 3 G C 5 Parental New New Parental Replicas

16 DNA Replication Making an exact duplicate of the DNA involves 30 different enzymes Begins at an origin of replication Helicase unwinds and unzips the DNA double helix (b) Replication forks 16

17 DNA Replication An RNA primer is synthesized at the origin of replication by Primase DNA polymerase III adds nucleotides in a 5 to 3 direction (b) Replication forks 17

18 DNA Replication DNA polymerase III adds nucleotides in a 5 to 3 direction Leading strand synthesized continuously in 5 to 3 direction Lagging strand synthesized 5 to 3 in short segments; overall direction is 3 to 5 (b) Replication forks 18

19 DNA Replication DNA polymerase I removes the RNA primers and replaces them with DNA When replication forks meet, ligases link the DNA fragments along the lagging strand Separation of the daughter molecules is complete (b) Replication forks 19

20 Overall Bacterial DNA Replication 4.Before synthesis of the lagging strand can start, a primase first constructs a short RNA primer to direct the DNA polymerase III. Synthesis can proceed only in short sections and produces segments of RNA primer and new DNA called Okazaki fragments. 3.The template for the lagging strand runs the opposite direction (3 to 5 ) and must be replicated backwards away from the replication fork so the DNA polymerase can add the nucleotides in the necessary 5 to 3 arrangement. 5. A second polymerase (DNA polymerase I) acts on the Okazaki fragments by removing the primers Open spaces in the lagging strand are filled in by a ligase that adds the correct nucleotides (a) Forks Lagging strand synthesis Nick 1. The chromosome tobe replicated is continuously unwound by a helicase, forming a replication fork with two template strands The template for the leading strand (bottom) is correctly oriented for the DNA polymerase III to add nucleotides in the 5 to 3 direction towards the replication fork, so it can be synthesized as a continuous strand. Note that direction of synthesis refers to the order of the new strand (red). Lagging strand synthesis Key: Template strand New strand RNA primer Helicase Primase DNA polymerase III DNA polymerase I Ligase Daughter cell Daughter cell (b)

21 Enzymes Involved in Replication 21

22 Concept Check: Why must the lagging strand of DNA be replicated in short pieces? A. Because of limited space B. Otherwise, the helix will become distorted C. The DNA polymerase can synthesize in only one direction D. To make proofreading of the code easier

23 Applications of the DNA code (a) (b) Information stored on the DNA molecule is conveyed to RNA molecules through the process of transcription Transcription of DNA DNA DSRNA SSRNA Regulatory RNAs trna mrna rrna The information contained in the RNA molecule is then used to produce proteins in the process of translation Ribosome (rrna+protein Translation of RNA Protein trna mrna Micro RNA, interfering RNA, antisense RNA, and riboswitches regulate transcription and translation Expression of DNA for structures and functions of cell 23

24 Gene-Protein Connection 1. Each triplet of nucleotides on the RNA specifies a particular amino acid 2. A protein s primary structure determines its shape and function 3. Proteins determine phenotype. Living things are what their proteins make them. 4. DNA is mainly a blueprint that tells the cell which kinds of proteins to make and how to make them DNA mrna (copy of one strand) Amino acids Triplets 1 Codon Single nucleotide Variations in the order and types will dictate the shape 24 and function of the protein

25 RNAs Single-stranded molecule made of nucleotides 5 carbon sugar is ribose 4 nitrogen bases adenine, uracil, guanine, cytosine Phosphate 25

26 RNA 3 types of RNA: Messenger RNA (mrna) carries DNA message through complementary copy; message is in triplets called codons (a) Messenger RNA (mrna) Ashort piece of messenger RNA (mrna illustrates the general structure of RNA: single strandedness, repeating phosphate-ribose sugar backbone attached to single nitrogen bases; use of uracil instead of thymine. A U G C U G A C U P P P P P P P P Codon 1 Codon 2 Codon 3 P = Phosphate R = Ribose U = Uracil Transfer RNA (trna) Ribosomal RNA (rrna) 26

27 RNA 3 types of RNA: Messenger RNA (mrna) Transfer RNA (trna) made from DNA; secondary structure creates loops; bottom loop exposes a triplet of nucleotides called anticodon which designates specificity and complements mrna; carries specific amino acids to ribosomes (b) Transfer RNA (trna) Left : The trna stand loops back on itself to form intrachain hydrogen bonds. The result is a cloverleaf structure, shown here in simplified form. At its bottom is an anticodon that specifies the attachment of a particular amino acid at the 39 end right A three-dimensional view of trna structure. G H bonds A G G A G G G A 5 A 3 Amino acid attachment site G C C G G C G A A A G A A A G G A G A A G A G G A G Hairpin loops Anticodon Anticodon Amino acid attachment site 5 3 Ribosomal RNA (rrna) 27

28 RNA 3 types of RNA: Messenger RNA (mrna) Transfer RNA (trna) Ribosomal RNA (rrna) component of ribosomes where protein synthesis occurs Amino acids Large subunit Exit site P A E Small transcript 5 trnas mrna transcript 28

29 Transcription: The First Stage of Gene Expression 1. RNA polymerase binds to promoter region upstream of the gene 2. RNA polymerase adds nucleotides complementary to the template strand of a segment of DNA in the 5 to 3 direction 3. Uracil is placed as adenine s complement 4. At termination, RNA polymerase recognizes signals and releases the transcript 100-1,200 bases long 29

30 Transcription Each gene contains a specific promoter region and a leader sequence for guiding the beginning of transcription. Next is the region of the gene that codes for a polypeptide and ends with a series of terminal sequences that stop translation. DNA is unwound at the promoter by RNA polymerase. Only one strand of DNA, called the template strand, is copied by the RNA polymerase. This strand runs in the 3' to 59 direction. The RNA polymerase moves along the strand, adding complementary nucleotides as dictated by the DNA template. The mrna strand reads in the 5' to 39 direction. RNA polymerase 5' Promoter region T A Initiation codon A T C G T emplate strand 5 Nontemplate strand Direction of transcription RNA polymerase binding site Leader sequence 3' 5' 3 3' G C A T C G T A Unwinding of DNA Nucleotide pool Termination sequences ( ) G A T G C C T A C G ( ) Intervening sequence of variable size T ermination sequence 4 The polymerase continues transcribing until it reaches a termination site, and the mrna transcript is released to be translated. Note that the section of the transcribed DNA is rewound into its original configuration. 5' Early mrna transcript Late mrna transcript Elongation 30

31 Translation: The Second Stage of Gene Expression All the elements needed to synthesize protein are brought together on the ribosomes Exit site Amino acids E P A Large subunit The process occurs in five stages: initiation, elongation, termination, and protein folding and processing 5 trnas Small transcript mrna transcript 31

32 First Base Position Third Base Position The Master Genetic Code Represented by the mrna codons and the amino acids they specify Code is universal among organisms Code is redundant U C A G } Second Base Position U C A G UUU UCU UAU UGU U Phenylalanine T yrosine Cysteine UUC UCC UAC UGC C Serine UUA UCA UAA UGA STOP** A Leucine } STOP** UUG } UCG UAG UGG Tryptophan G CUU CCU CAU Histidine CGU U CUC CCC CAC CGC C Leucine Proline Arginine CUA CCA CAA CGA A Glutamine } CUG CCG CAG CGG G AUU Isoleucine ACU AAU Asparagine AGU Serine U AUC ACC AAC AGC C Threonine AUA AC A AAA AGA A AUG* Methionine ACG AAG } Lysine Arginine START AGG } G GUU GCU GAU GGU U Aspartic acid GUC GCC GAC GGC C Valine Alanine Glycine GUA GC A GAC GGA A Glutamic acid GUG GCG GAG GGG G *This codon initiates translation. **For these codons, which give the orders to stop translation, there are no corresponding trnas with amino acids. } } } } } } } 32

33 Interpreting the DNA Code Transcription produces an mrna complementary to the DNA gene DNA triplets mrna codons Nontemplate strand Template strand During translation, trnas use their anticodon to interpret the mrna codons and bring in the amino acids trna anticodons Protein (amino acid specified) UAC F-Methionine GAC Leucine UGA Threonine UGC Threonine Same amino acid; has a different codon and anticodon 33

34 Translation Ribosomes assemble on the 5 end of an mrna transcript Ribosome scans the mrna until it reaches the start codon, usually AUG Exit site Amino acids E P A Large subunit A trna molecule with the complementary anticodon and methionine amino acid enters the P site of the ribosome and binds to the mrna 5 trnas 34 Small transcript mrna transcript

35 Translation A second trna with the complementary anticodon fills the A site f Met Leucine Anticodion m RNA Start codon Entrance of trnas 1 and 2 CCG 35

36 Translation A peptide bond is formed is formed between the amino acids on the neighboring trnas Peptide bond 1 CCG Fermationof peptide bond 36

37 Translation The first trna is released and the ribosome slides down to the next codon Empty trna UAC CCG P site Discharge of trna 1 at E site 37

38 Translation Another trna fills the A site and a peptide bond is formed Proline Peptide bond 2 A A 2 First translocation: trna 2 shifts into p site ; trna 3 enters ribosome at A UAG Formation of peptide bond 38 UAG

39 Translation This process continues until a stop codon is reached Alanine Peptide bond 3 A 3 G G C AUC AUC Discharge of trna 2; second translocation; trna 4 enters ribosome Formation of peptide bond Stop codon

40 Translation Termination Termination codons UAA, UAG, and UGA are codons for which there is no corresponding trna When this codon is reached, the ribosome falls off and the last trna is removed from the polypeptide 40

41 First Base Position Third Base Position U C A G } Second Base Position U C A G UUU UCU UAU UGU U Phenylalanine T yrosine Cysteine UUC UCC UAC UGC C Serine UUA UCA UAA UGA STOP** A Leucine } STOP** UUG } UCG UAG UGG Tryptophan G CUU CCU CAU Histidine CGU U CUC CCC CAC CGC C Leucine Proline Arginine CUA CCA CAA CGA A Glutamine } CUG CCG CAG CGG G AUU Isoleucine ACU AAU Asparagine AGU Serine U AUC ACC AAC AGC C Threonine AUA AC A AAA AGA A AUG* Methionine ACG AAG } Lysine Arginine START AGG } G GUU GCU GAU GGU U Aspartic acid GUC GCC GAC GGC C Valine Alanine Glycine GUA GC A GAC GGA A Glutamic acid GUG GCG GAG GGG G *This codon initiates translation. **For these codons, which give the orders to stop translation, there are no corresponding trnas with amino acids. } } } } } } } 41 41

42 Concept Check: Transfer RNA is the molecule that A. contributes to the structure of ribosomes B. adapts the genetic code to protein structure C. transfers the DNA code to mrna D. provides the master code for amino acids

43 Polyribosomal Complex Polyribosomal complex allows for the synthesis of many protein molecules simultaneously from the same mrna molecule. mrna RNA polymerase Transcription Start of translation mrna (a) Ribosomes Growing polypeptides 1 Polypeptide Polyribosomal complex Start (c) Steven McKnight and Oscar L Mille, Department of Biolog, University of virginia 43 (b)

44 Eukaryotic Transcription and Translation 1. Do not occur simultaneously transcription occurs in the nucleus and translation occurs in the cytoplasm 2. Eukaryotic start codon is AUG, but it does not use formyl-methionine 3. Eukaryotic mrna encodes a single protein, unlike bacterial mrna which encodes many 4. Eukaryotic DNA contains introns intervening sequences of noncoding DNA which have to be spliced out of the final mrna transcript 44

45 Splicing of Eukaryotic pre-mrna Removal of introns and connection of exons DNA template Primary mrna transcript E II E E I E Exon Intron E II E E I E Does not occur in Prokaryotes Occurs In nucleus Transcript processed by special enzymes Larat forming Spliceosomes E E E E Lariat excised Spliceosomes released Exons spliced together E E E E Occurs in cytoplasm mrna transcript can now be translated 45

46 Regulation of Protein Synthesis and Metabolism Genes are regulated to be active only when their products are required In prokaryotes this regulation is coordinated by operons, a set of genes, all of which are regulated as a single unit 46

47 Operons 2 types of operons: Inducible operon is turned ON by substrate: catabolic operons - enzymes needed to metabolize a nutrient are produced when needed Repressible genes in a series are turned OFF by the product synthesized; anabolic operon enzymes used to synthesize an amino acid stop being produced when they are not needed 47

48 Lactose Operon: Inducible Operon Made of 3 segments: 1. Regulator gene that codes for repressor 2. Control locus composed of promoter and operator 3. Structural locus made of 3 genes each coding for an enzyme needed to catabolize lactose b-galactosidase hydrolyzes lactose permease brings lactose across cell membrane b-galactosidase transacetylase uncertain function Repressor RNA polymerase 1 Operon Off In the absence of lactose, a repressor protein (the product of a regulatory gene located elsewhere on the bacterial chromosome) attaches to the operator of the operon. This e ffectively locks the operator and prevents any transcription of structural genes downstream (to its right). Suppression of transcription (and consequentl, of translation) prevents the unnecessary synthesis of enzymes for processing lactose. Locked Translation 48

49 Lac Operon Normally off In the absence of lactose, the repressor binds with the operator locus and blocks transcription of downstream structural genes Repressor RNA polymerase 1 Operon Off In the absence of lactose, a repressor protein (the product of a regulatory gene located elsewhere on the bacterial chromosome) attaches to the operator of the operon. This e ffectively locks the operator and prevents any transcription of structural genes downstream (to its right). Suppression of transcription (and consequentl, of translation) prevents the unnecessary synthesis of enzymes for processing lactose. Locked Translation 49

50 Lac Operon Lactose turns the operon on by acting as the inducer Binding of lactose to the repressor protein changes its shape and causes it to fall off the operator. RNA polymerase can bind to the promoter. Structural genes are transcribed. 2 Operon On Upon entering the cell, the substrate (lactose) becomes a genetic inducer by attaching to the represso, which loses its grip and falls away. The RNA polymerase is now free to bind to the promoter and initiate transcription, and the enzymes produced by translation of the mrna perform the necessary reactions on their lactose substrate. Inactive repressor RNA polymerase active Lactose (inducer) mrna TRanslation into enzymes Lactose transported, digested, and used In metabolism 50

51 Arginine Operon: Repressible Normally on and will be turned off when the product of the pathway is no longer required RNA polymerase 1 Operon On A repressible operon remains on when its nutrient products (here, arginine) are in great demand by the cell. The repressor cannot bind to the operator at low nutrient levels. mrna Repressor is inactive (wrong shape to attach to operator) Rep Enzymes synthesize arginine. Arginine is immediately used in metabolism 51

52 Arginine Operon: Repressible When excess arginine is present, it binds to the repressor and changes it. Then the repressor binds to the operator and blocks arginine synthesis. Arginine is the corepressor. (1) Repressor is active (correct shape achieved) RNA polymerase Arginine accumulates 2 Operon Off The operon is repressed when (1) arginine builds up and, serving as a corepressor. activates the repressor.(2) The repressor complex affixes to the operator and blocks the RNA A polymerase and further transcription of genes for arginine synthesis. (2) 52

53 Concept Check: If an operon s repressor is in its active form that means A. Transcription from the operon is occurring B. Transcription from the operon is not occurring

54 Mutations: Changes in the Genetic Code A change in phenotype due to a change in genotype (nitrogen base sequence of DNA) is called a mutation A natural, nonmutated characteristic is known as a wild type (wild strain) An organism that has a mutation is a mutant strain, showing variance in morphology, nutritional characteristics, genetic control mechanisms, resistance to chemicals, etc. 54

55 Isolating Mutants (a) Treatment of culture with a mutagen. Replica block Replica Plating technique allows identification of mutants (b) Inoculate a plate containing complete growth medium and incubate. Both wild-type and mutants form colonies. (c) Velvet surface (sterilized) Master plate (complete medium) (c) Replica plate (complete medium) Replica plate (medium minus nutrient) Incubation (d) All strains grow No colony Mutant present colonies do not grow Mutant colony can be located and isolated 55

56 Causes of Mutations Spontaneous mutations random change in the DNA due to errors in replication that occur without known cause Induced mutations result from exposure to known mutagens, physical (primarily radiation) or chemical agents that interact with DNA in a disruptive manner 56

57 Categories of Mutations Point mutation addition, deletion, or substitution of a few bases Missense mutation causes change in a single amino acid Nonsense mutation changes a normal codon into a stop codon Silent mutation alters a base but does not change the amino acid Back-mutation when a mutated gene reverses to its original base composition Frameshift mutation when the reading frame of the mrna is altered 57

58 Effect of Mutations 58

59 Repair of Mutations Since mutations can be potentially fatal, the cell has several enzymatic repair mechanisms in place to find and repair damaged DNA DNA polymerase proofreads nucleotides during DNA replication Mismatch repair locates and repairs mismatched nitrogen bases that were not repaired by DNA polymerase Light repair for UV light damage Excision repair locates and repairs incorrect sequence by removing a segment of the DNA and then adding the correct nucleotides 59

60 Excision Repair Mechanism Copyright The McGraw-Hill Companies, Inc. Permission required for reproduction or display. Enzyme complex I Removed (a) Enzyme complex II Added (b) 60 (c)

61 The Ames Test Culture of Salmonella bacteria, histidine (-) Any chemical capable of mutating bacterial DNA can similarly mutate mammalian DNA Agricultural, industrial, and medicinal compounds are screened using the Ames test Indicator organism is a mutant strain of Salmonella typhimurium that has lost the ability to synthesize histidine This mutation is highly susceptible to back-mutation In the control setup, bacteria are plated on a histidine-free medium containing liver enzymes but lacking the test agent. (a) Control Plate Minimal medium lacking histidine and test chemical his( + ) colonies arising from spontaneous back-mutation Incubation (12 h) Any colonies that form have back-mutated to his( + ) The experimental plate is prepared the same way except that it contains the test agent. (b) Test Plate Minimal medium with test chemical and no histidine his( + ) colonies in presence of the chemical (c) The degree of mutagenicity of the chemical agent can be calculated by comparing the number of colonies growing on the control plate with the number on the test plate. Chemicals that induce an increased incidence of back-mutation (right side) are considered carcinogens.

62 Positive and Negative Effects of Mutations Mutations leading to nonfunctional proteins are harmful, possibly fatal Organisms with mutations that are beneficial in their environment can readily adapt, survive, and reproduce these mutations are the basis of change in populations Any change that confers an advantage during selection pressure will be retained by the population 62

63 Concept Check: Which of the following mutations would cause a frameshift mutation? A. Silent mutation B. Missense mutation C. Nonsense mutation D. Deletion mutation

64 DNA Recombination Events Genetic recombination occurs when an organism acquires and expresses genes that originated in another organism 3 means for genetic recombination in bacteria: 1. Conjugation 2. Transformation 3. Transduction 64

65 Conjugation Conjugation transfer of a plasmid or chromosomal fragment from a donor cell to a recipient cell via a direct connection Gram-negative cell donor has a fertility plasmid (F plasmid, F factor) that allows the synthesis of a conjugative pilus Recipient cell is a related species or genus without a fertility plasmid Donor transfers fertility plasmid through pilus F factor 1 Physical Conjugation Bacterial chromosome F + Pilus Sex pilus makes contact with F recipient cell F + Sex pilus contracts, bringing cells together. F + F The pilus of donor cell (top) attaches to receptor on recipient cell and retracts to draw the two cells together. This is the mechanism for gram negative bacteria. 65

66 Conjugation High-frequency recombination donor s fertility plasmid has been integrated into the bacterial chromosome F factor (plasmid) F Factor Transfer Bridge made with pilus Donor Recipient F + F Chromosome F + F Hfr cell Donor Hfr Transfer Recipient F factor Integration of F facter into choromosome Pilus Chromosome Partial copy of donor chromosome When conjugation occurs, a portion of the chromosome and a portion of the fertility plasmid are transferred to the recipient F factor being copied F + F F factor 2 Transfer of the F facter, or 3 conjugative plasmid Bridge broken Donated genes High-frequency (Hfr) transfer involves transmission of chromosomal genes from a donor cell to a recipient cell. The donor chromosome is duplicated and transmitted in part to a recipient cell, where it is integrated into the chromosome.

67 Transformation Transformation chromosome fragments from a lysed cell are accepted by a recipient cell; the genetic code of the DNA fragment is acquired by the recipient Donor and recipient cells can be unrelated Useful tool in recombinant DNA technology DNA transport system Receptor Cap + Cap + Cap + ds DNA fragment (blue) with new gene (red) binds to a surface receptor on a competent recipient cell. DNA is converted to one strand and transported into the cell, by the DNA transport system. The DNA strand aligns itself with a compatible region on the recipient chromosome. The DNA strand is incorporated into the recipient chromosome Transformed cell Cap + Recipient is now transformed with gene for synthesizing a capsule. 67

68 Griffith s Work on Transformation Strain of Colony Cell Type Capsule Effect Smooth (S) Live S strain (c) Heat-killed S strain Survives (a) Dies Live R strain No capsule Rough (R) (b) Live R strain Survives (d) Heat-killed S strain Live S and R strains isolated from dead mouse Dies 68

69 Transduction Transduction bacteriophage serves as a carrier of DNA from a donor cell to a recipient cell Two types: Generalized transduction random fragments of disintegrating host DNA are picked up by the phage during assembly; any gene can be transmitted this way Specialized transduction a highly specific part of the host genome is regularly incorporated into the virus 69

70 Transduction Phage DN A (1) Donor (host) chromosome (1) Prophage within the bacterial chromosome Generalized transduction (2) Cell A (donor) Parts of phage Separated piece of host DNA (2) Excised phage DNA contains some bacterial DNA Specialized transduction (3) Newly assembled phage incorporating piece of host DNA (3) New viral particles are synthesized Lysis (4) (4) DNA from donor (5) Infection of recipient cell transfers bacterial DNA to a new cell Cell B (ecipient) (6) Incorporated into chromosome (5) Cell survives and utilizes transduced DNA Recombination results in two possible outcomes. 70

71 Intermicrobial DNA Exchange 71

72 Transposons Special DNA segments that have the capability of moving from one location in the genome to another jumping genes Cause rearrangement of the genetic material (1) Can move from one chromosome site to another, from a chromosome to a plasmid, or from a plasmid to a chromosome (2) (3) (4) May be beneficial or harmful 72

73 Genetics of Animal Viruses Viral genome - one or more pieces of DNA or RNA; contains only genes needed for production of new viruses Requires access to host cell s genetics and metabolic machinery to instruct the host cell to synthesize new viral particles 73

74 Multiplication of Double-Stranded DNA Viruses Viral proteins Host cell cytoplasm Viral DNA Nuclear pore Viral mrna Nucleus 5 Mature virus Replicated viral DNA 6 Host DNA 74

75 Multiplication of Positive-Strand, Single-Stranded RNA Viruses Virus (+) 1 Viral RNA (+) 2 ( ) 3 4 Viral proteins 5 Capsid Cytoplasm Nucleus 75

76 Concept Check: An F+ Cell A. Has undergone Conjugation B. Can undergo Conjugation C. Has undergone Transformation D. Can undergo Transformation E. Has undergone Transduction F. Can undergo Transduction