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1 The world leader in serving science
TaqMan® Drug Metabolism Genotyping Assays on OpenArray® Plates June 28, 2014
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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.
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CYP2D6*2A g.-1584C>G, rs1080985 C__32407252_30
Plasmid Samples
gDNA Samples
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.
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CYP2D6 g.2850C>T , rs16947 C__27102425_10
Plasmid Samples
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.
gDNA Samples
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CYP2D6*2 g.4180G>C, rs1135840 C__27102414_10
Plasmid Samples
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.
gDNA Samples
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CYP2D6*4 g.1846G>A , rs3892097 C__27102431_D0
Plasmid Samples
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.
gDNA Samples
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CYP2D6*7 g.2935A>C, rs5030867 C__32388575_A0 – recommended *7 assay version
Plasmid Samples
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.
gDNA Samples
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CYP2D6*7 g.2935A>C , rs5030867 C__32388575_30 - improved assay version is available*
Plasmid Samples
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.
gDNA Samples
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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.
gDNA Samples
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CYP2D6*8 g.1758G>T, rs5030865 C_30634117C_20 - improved assay version is available*
Plasmid Samples
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.
gDNA Samples
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CYP2D6*9 g.2613_2615delAGA, rs28371720 C__32407229_60
Plasmid Samples
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.
gDNA Samples
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CYP2D6*10 g.100C>T, rs1065852 C__11484460_40
Plasmid Samples 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.
gDNA Samples
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CYP2D6*11 g.883G>C, rs5030863 C__30634118_A0
Plasmid Samples
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.
gDNA Samples
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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.
gDNA Samples
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CYP2D6*14 g.1758G>A , rs5030865 C_30634117D_30 - improved assay version is available*
Plasmid Samples
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.
gDNA Samples
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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.
gDNA Samples
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CYP2D6*17 g.1023C>T, rs28371706 C___2222771_40 - improved assay version is available*
Plasmid Samples
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.
gDNA Samples
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CYP2D6*35 g.31G>A, rs769258 C__27102444_80
Plasmid Samples gDNA Sample Cycle 30
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.
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CYP2C9*3/*18 c.1075A>C, g.42614A>C, rs1057910 C__27104892_10
Plasmid Samples 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’.
gDNA Samples
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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’.
gDNA Samples
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CYP2C19*2 g.19154G>A. c.681G>A (splicing defect) , rs4244285 C__25986767_70
Plasmid Samples
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’.
gDNA Samples
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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’.
gDNA Samples
108
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).
gDNA Samples
109
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.
gDNA Samples