TaqMan Drug Metabolism Genotyping Assays on OpenArray Plates June 28, 2014 The world leader in serving science 1
TaqMan Drug Metabolism Genotyping Assays on OpenArray Plates Life Technologies has pretested over 300 TaqMan DME and SNP assays to highly studied, important DME and other gene variants on OpenArray plates run on the QuantStudio 12K Flex Real-Time PCR System. Assays were tested on this system with 44 Coriell gdnas (22 each African American and Caucasian samples, a subset of the TaqMan DME Assay 90 sample validation panel) and most were also tested with synthetic plasmid constructs representing each genotype. You can download the PGx Common Markers file that contains a list of the most commonly requested assays from this set at: www.lifetechnologies.com/pgx The file includes common allele names, context sequences, and other useful annotations. This PowerPoint document contains screen shots of the test data for the most commonly requested PGx and clinical research target assays. For Research Use Only. Not for use in diagnostic procedures. 2014 Thermo Fisher Scientific. All rights reserved. All trademarks are the property of Thermo Fisher Scientific and its subsidiaries unless otherwise specified. 2
CYP2D6*2A g.-1584c>g, rs1080985 C 32407252_30 The CYP2D6*2 allele can be duplicated. Samples having 3 copies of CYP2D6 that are heterozygous for -1584C>G may run within or beside the heterozygous cluster. 3
CYP2D6 g.2850c>t, rs16947 C 27102425_10 2850C>T is a common SNP that is found in *2 and many other CYP2D6 star alleles. The *2 allele can be duplicated. Samples having 3 copies of CYP2D6 that are heterozygous for 2850C>T may run within or beside the heterozygous cluster. 4
CYP2D6*2 g.4180g>c, rs1135840 C 27102414_10 4180G>C is a common SNP that is found in *2 and many other CYP2D6 star alleles. The *2 allele can be duplicated. Samples having 3 copies of CYP2D6 that are heterozygous for 4180G>C may run within or beside the heterozygous cluster. 5
CYP2D6*3 g.2549dela, rs35742686 C 32407232_50 6
CYP2D6*4 g.1846g>a, rs3892097 C 27102431_D0 The CYP2D6 *4 allele can be duplicated. Samples having 3 copies of CYP2D6 that are heterozygous for 1846G>A may run within or beside the heterozygous cluster. 7
CYP2D6*6 g.1707delt, rs5030655 C 32407243_20 8
CYP2D6*7 g.2935a>c, rs5030867 C 32388575_A0 recommended *7 assay version C 32388575_A0 is an improved version of C 32388575_30 (slide 10). It has higher amplification signal due to reduction of NTC signal. We recommend using C 32388575_A0 to interrogate CYP2D6*7 g.2935a>c. 9
CYP2D6*7 g.2935a>c, rs5030867 C 32388575_30 - improved assay version is available* The high NTC signal seen with C 32388575_30 is due to probe cleavage in the absence of template and amplification. It does not interfere with genotyping unless sample the amplification signal is low (e.g. due to low sample quantity). *We recommend using the improved assay design: C 32388575_A0 (slide 9). It has higher amplification signal due to reduction of NTC signal. 10
CYP2D6*8 g.1758g>t, rs5030865 C_30634117C_K0 recommended *8 assay version C_30634117C_K0 reports the major (G) and one minor (T) allele (CYP2D6*8) of a triallelic SNP. C_30634117D_M0 reports the major (G) and other minor (A) allele (CYP2D6*14). Both assays should be independently run on samples and data analyzed as described in the PGx Experiments User Guide Chapter 5 section on triallelic and adjacent SNPs. C_30634117C_K0 is an improved version of C_30634117C_20 (slide 12), which has higher amplification signal due to a decrease in amplicon size. 11
CYP2D6*8 g.1758g>t, rs5030865 C_30634117C_20 - improved assay version is available* C_30634117C_20 reports the major (G) and one minor (T) allele (CYP2D6*8) of a triallelic SNP. C_30634117D_30 reports the major (G) and other minor (A) allele (CYP2D6*14). Both assays should be independently run on samples and data analyzed as described in the PGx Experiments User Guide Chapter 5 section on triallelic and adjacent SNPs. * We recommend using the improved assay design: C_30634117C_K0 (slide 11). It has higher amplification signal due to a decrease in amplicon size. 12
CYP2D6*9 g.2613_2615delaga, rs28371720 C 32407229_60 C 32407229_60 targets the CYP2D6*9 deletion variant, which can be denoted as g.2613_2615delaga or g.2615_2617delaag. The nucleotide deletion results in deletion of amino acid K281. 13
CYP2D6*10 g.100c>t, rs1065852 C 11484460_40 Cycle 27 The CYP2D6*10 allele can be duplicated. Samples having 3 copies of CYP2D6 that are heterozygous for 100C>T may run within or beside the heterozygous cluster. 14
CYP2D6*11 g.883g>c, rs5030863 C 30634118_A0 C 30634118_A0 specifically amplifies CYP2D6 by targeting base differences between CYP2D6 and CYP2D7. Two of these base differences occur as SNPs in some CYP2D6*4 alleles, which thus cannot be amplified by the *11 assay. The CYP2D6*4 C 27102431_D0 assay should be run on the same samples for complete genotype analysis, as *4/*4 samples will not amplify with the *11 assay, and samples that run as *11/*11 may have *11/*11 or *4/*11 genotypes, which can be discerned by the *4 assay results. 15
CYP2D6*12 g.124g>a, rs5030862 C 27531552_A0 16
CYP2D6*14 g.1758g>a, rs5030865 C_30634117D_M0 recommended *14 assay version C_30634117D_M0 reports the major (G) and one minor (A) allele (CYP2D6*14) of a triallelic SNP. C_30634117C_K0 reports the major (G) and other minor (T) allele (CYP2D6*8). Both assays should be independently run on samples and data analyzed as described in the PGx Experiments User Guide Chapter 5 section on triallelic and adjacent SNPs. C_30634117D_M0 is an improved version of C_30634117D_30 (slide 18), which has higher amplification signal due to a decrease in amplicon size. 17
CYP2D6*14 g.1758g>a, rs5030865 C_30634117D_30 - improved assay version is available* C_30634117D_30 reports the major (G) and one minor (A) allele (CYP2D6*14) of a triallelic SNP. C_30634117C_20 reports the major (G) and other minor (T) allele (CYP2D6*8). Both assays should be independently run on samples and data analyzed as described in the PGx Experiments User Guide Chapter 5 section on triallelic and adjacent SNPs. * We recommend using the improved assay design: C_30634117D_M0 (slide 17). It has higher amplification signal due to a decrease in amplicon size. 18
CYP2D6*15 g.137-138inst, rs72549357 C 32407245_40 19
CYP2D6*17 g.1023c>t, rs28371706 C 2222771_A0 recommended *14 assay version The CYP2D6*17 allele can be duplicated. Samples having 3 copies of CYP2D6 that are heterozygous for 1023C>T may run within or beside the heterozygous cluster. C 2222771_A0 is an improved version of C 2222771_40 (slide 21). It has been observed that C 2222771_40 cannot amplify a subset of *4 alleles and thus some *4/*17 samples will run as *17 homozygotes. E.g. In the experiment shown, samples NA17116 and NA17121 run as heterozygotes with C 2222771_A0 whereas they run as *17/*17 homozygotes with C 2222771_40. 20
CYP2D6*17 g.1023c>t, rs28371706 C 2222771_40 - improved assay version is available* The CYP2D6*17 allele can be duplicated. Samples having 3 copies of CYP2D6 that are heterozygous for 1023C>T may run within or beside the heterozygous cluster. It has been observed that C 2222771_40 cannot amplify a subset of *4 alleles and thus some *4/*17 samples will run as *17 homozygotes. E.g. In the experiment shown, samples NA17116 and NA17121 run as homozygotes with C 2222771_40 whereas they run as *17 heterozygotes with C 2222771_A0. *We recommend using the improved assay: C 2222771_A0 (slide 20) to interrogate CYP2D6*17 g.1023c>t. 21
CYP2D6*29 g 3183G>A, rs59421388 C 34816113_20 22
CYP2D6*35 g.31g>a, rs769258 C 27102444_80 Cycle 30 gdna Sample The CYP2D6*35 allele can be duplicated. Samples having 3 copies of CYP2D6 that are heterozygous for 31G>A may run within or beside the heterozygous cluster. 23
CYP2D6*41 g.2988g>a (splicing defect), rs28371725 C 34816116_20 24
CYP2C9*2 c.430c>t g.3608c>t, rs1799853 C 25625805_10 25
CYP2C9*3/*18 c.1075a>c, g.42614a>c, rs1057910 C 27104892_10 Cycle 32 The polymorphic CYP2C9*3/*18 c.1075a>c SNP is adjacent to the rare CYP2C9*4 c.1076t>c SNP that is detected by C 30634131_20 (slide 27). The *3/*18 and *4 SNP assays can be used to detect the 3 possible haplotypes described for this locus (alleles *3/*18 1076C and *4 1075C do not occur together in a haplotype). Analyze data as described in the PGx Experiments User Guide Chapter 5 section TaqMan DME genotyping assays to triallelic SNPs and adjacent SNP targets. 26
CYP2C9*4 c. 1076T>C, g.42615t>c, rs56165452 C 30634131_20 Note: the samples with higher signal are wild type for the adjacent *3/*18 c.1075a>c SNP, those in the lower split cluster are heterozygous for this SNP, and the sample that does not amplify is homozygous for the 1075C allele. The rare CYP2C9*4 c.1076t>c SNP is adjacent to the polymorphic CYP2C9*3/*18 c.1075a>c SNP, which is detected by C 27104892_10 (slide 26). The *3/*18 and *4 SNP assays can be used to detect the 3 possible haplotypes described for this locus (alleles *3/*18 1076C and *4 1075C do not occur together in a haplotype). Analyze data as described in the PGx Experiments User Guide Chapter 5 section TaqMan DME genotyping assays to triallelic SNPs and adjacent SNP targets. 27
CYP2C9*5 c.1080c>g g.42619c>g, rs28371686 C 27859817_40 Cycle 27 28
CYP2C9*6 c.818dela g.10601dela C 32287221_20 29
CYP2C9*8 c.449g>a g.3627g>a, rs7900194 C 25625804_10 30
CYP2C9*11 c.1003c>t g.42542c>t, rs28371685 C 30634132_70 31
CYP2C9*13 c.269t>c g.3276t>c, rs72558187 C 34816136_20 32
CYP2C9*27 c.449g>t g.3627g>t, rs7900194 C_25625804D_20 33
CYP2C19*2 g.19154g>a. c.681g>a (splicing defect), rs4244285 C 25986767_70 The polymorphic CYP2C19*2 c.681g>a SNP is adjacent to the rare CYP2C19*10 c.680c>t SNP that is detected by C 30634128_10 (slide 35). The *2 and *10 SNP assays can be used to detect the 3 possible haplotypes described for this locus (alleles *2 681A and *10 680T do not occur together in a haplotype). Analyze data as described in the PGx Experiments User Guide Chapter 5 section TaqMan DME genotyping assays to triallelic SNPs and adjacent SNP targets. 34
CYP2C19*10 c.680c>t g.19153c>t, rs6413438 C 30634128_10 The rare CYP2C19*10 c.680c>t SNP is adjacent to the polymorphic CYP2C19*2 c.681g>a SNP that is detected by C 25986767_70 (slide 34). The *2 and *10 SNP assays can be used to detect the 3 possible haplotypes described for this locus (alleles *2 681A and *10 680T do not occur together in a haplotype). Analyze data as described in the PGx Experiments User Guide Chapter 5 section TaqMan DME genotyping assays to triallelic SNPs and adjacent SNP targets. 35
CYP2C19*3 g.17948g>a, rs4986893 C 27861809_10 36
CYP2C19*4 c.1a>g g.1a>g, rs28399504 C 30634136_10 37
CYP2C19*5 g.90033c>t, rs56337013 C 27861810_10 38
CYP2C19*6 g.12748g>a, rs72552267 C 27531918_10 39
CYP2C19*7 g.19294t>a, rs72558186 C 30634127_10 40
CYP2C19*8 g.12711t>c, rs41291556 C 30634130_30 41
CYP2C19*9 c.431g>a g.12784g>a, rs28399507 C 25745302_30 42
CYP2C19*17 g.-806c>t, rs12248560 C 469857_10 43
CYP3A4*1B g.-392a>g, rs2740574 C 1837671_50 44
CYP3A4*2 g.15713t>c, rs55785340 C 30634204_10 45
CYP3A4*3 c.1334t>c, rs4986910 C 27535825_20 46
CYP3A4*12 c.1117 C>T g.21896c>t, rs12721629 C 30634202_10 47
CYP3A4*16 c.554c>g g.15603c>g, rs12721627 C 30634207_10 48
CYP3A4*17 c.566 T>C g.15615t>c, rs4987161 C 27859822_10 49
CYP3A4*22 g.15389c>t, rs35599367 C 59013445_10 50
CYP3A5*9 g.19386g>a, rs28383479 C 30633863_10 51
CYP3A5*3B g.3705c>t, rs28383468 C 30633871_50 52
CYP3A5*2 g.27289c>a, rs28365083 C 30633862_10 53
CYP3A5*3 g.6986a>g, rs776746 C 26201809_30 54
CYP3A5*3/*10 g.31611c>t, rs15524 C 8303531_40 55
CYP3A5*6 g.14690g>a, rs10264272 C 30203950_10 56
CYP3A5*7 g.27131_27132inst, rs41303343 C 32287188_10 57
CYP3A5*8 g.3699c>t, rs55817950 C 30633872_10 58
CYP1A2*1 g.-3860g>a, rs2069514 C 15859191_30 59
CYP1A2*1 g.-2467delt, rs35694136 C 60142977_10 60
CYP1A2*1K g.-729c>t, rs12720461 C 30634146_10 61
CYP1A2*1 g.-739t>g, rs2069526 C 16017734_10 62
CYP1A2*1F g.-163c>a, rs762551 C 8881221_40 Cycle 30 63
CYP1A2*4 g.2499a>t, rs72547516 C 30634246_10 64
CYP1A2*3 g.2116g>a, rs56276455 C 30634247_20 65
CYP1A2*6 g.5090c>t, rs28399424 C 30634244_20 66
CYP1A2*7 g.3533g>a, rs56107638 C 30634145_10 67
CYP1A2*8 g.5166g>a, rs72547517 C 72649745_10 68
CYP1A2*11 g.558c>a, rs72547513 C 34816151_10 69
CYP1A2*15 g.125c>g, rs72547511 C 72649743_10 70
CYP1A2*16 g.2473g>a, rs72547515 C 72649744_20 71
CYP1A2 g.5347t>c, rs2470890 C 1642455_10 72
CYP2B6 c.516g>t g.15631g>t, rs3745274 C 7817765_60 73
CYP2B6*5 c.1459c>t, g.25505c>t, rs3211371 C 30634242_40 74
CYP2B6*10 c.64c>t g.64c>t, rs8192709 C 2818162_20 Cycle 30 75
CYP2B6*16/*18 c.983t>c g.21011t>c, rs28399499 C 60732328_20 76
CYP2B6*22 g.-82t>c, rs34223104 C 27830964_10 77
CYP2C8*2 c.805a>t g.11054a>t, rs11572103 C 30634034_10 78
CYP2C8*3 c.1196a>g g.30411a>g, rs10509681 C 25625782_20 79
CYP2C8*4 c.792c>g g.11041c>g, rs1058930 C 25761568_20 80
COMT c.322g>a or c.472g>a, rs4680 C 25746809_50 81
DRD2*A1 32806C>T Taq1A, rs1800497 C 7486676_10 82
SLCO1B1*5 g.37041t>c, rs4149056 C 30633906_10 83
UGT1A1*6 c.211g>a g.211g>a, rs4148323 C 559715_20 84
UGT1A1*60 g. -3279T>G, rs4124874 C 1432134_10 85
VKORC1-prom, rs9923231 C 30403261_20 86
VKORC1 358C>T P83L, rs7200749 C 29057362_10 Cycle 30 87
VKORC1 497T>G, rs2884737 C 16147492_20 88
VKORC1 1173C>T, rs9934438 C 30204875_10 89
VKORC1 3730G>A, rs7294 C 7473918_10 90
Apo-ε2 c.526c>t, rs7412 C 904973_10 91
Apo-ε4 c.t388c, rs429358 C 3084793_20 92
HLA-B*1502 tag rs3745274, rs3909184 C 8692805_10 93
HTR2A -1438G>A, rs6311 C 8695278_10 94
HTR2C -759C>T, rs3813929 C 27488117_10 95
F2 G20210A, rs1799963 C 8726802_20 96
Factor V R506Q Leiden mutation, rs6025 C 11975250_10 97
F7 rs6042, rs6042 C 3046089_20 98
MTHFR C677T Ala222Val, rs1801133 C 1202883_20 99
MTHFR A1298C Glu429Ala, rs1801131 C 850486_20 100
OPRM1 A118G Asn40Asp, rs1799971 C 8950074_1_ 101
VKORC1-1639G>A, rs9923231 C 30403261_20 102
VKORC1 1173C>T, rs9934438 C 30204875_10 103
VKORC1 3730G>A, rs7294 C 7473918_10 104
VKORC1 1542G>C, rs8050894 C 2847860_10 105
VKORC1 2255C>T, rs2359612 C 26291751_10 106
VKORC1 6009C>T, rs17708472 C 32928084_10 107
Gender Assay AMEL_Y-FAM_X-VIC, hcv990000001 C_990000001_10 C_990000001_10, targets a gender-specific polymorphic region in the amelogenin gene. A 6 base deletion occurring in the X-specific AMEL gene is detected by the VIC dye probe, whereas the FAM dye probe detects Y-specific sequences. Male samples run in the heterozygous cluster position and female samples run in the VIC homozygous cluster. Note: some males lack the Y-specific amelogenin gene and will type as female. The C_990000001_10 assay should be run in combination with a Y-chromosome assay to identify any mistyped samples (e.g. C 8938211_20; see slide 109). 108
Y-chr assay, rs3913290 C 8938211_20 This Y-chromosome-specific assay will amplify only male samples; female samples run with or close to the NTCs. 109
Legal notification For Research Use Only. Not for use in diagnostic procedures. 2014 Thermo Fisher Scientific. All rights reserved. All trademarks are the property of Thermo Fisher Scientific and its subsidiaries unless otherwise specified. 110