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TotalPlex gene amplification using bulging primers for pharmacogenetic analysis of acute lymphoblastic leukemia.

Abstract：

:Genetic polymorphism among patients with acute lymphoblastic leukemia (ALL) is an important factor in the effectiveness and toxicity of anti-leukemic drugs. Genotyping of various polymorphisms that impact the outcome of anti-leukemic drug therapy (pharmacogenetics) presents an attractive approach for developing individualized therapy. We developed an easy and accurate method of analyzing multiple genes using a small amount of DNA, which we termed TotalPlex amplification. We used 16 pairs of specific bulging specific primers (SBS primers) for simultaneous amplification of 16 loci in a single PCR tube. Sixteen single nucleotide polymorphisms (SNPs) (CYP3A4*1B A>G, CYP3A5*3 G>A, GSTP1 313 A>G, GSTM1 deletion, GSTT1 deletion, MDR1 exon 21 G>T/A, MDR1 exon 26 C>T, MTHFR 677 C>T, MTHFR 1298 A>C, NR3C1 1088 A>G, RFC 80 G>A, TPMT 238 G>C, TPMT 460 G>A, TPMT 719 A>G, VDR intron 8 G>A, VDR FokI T>C) that have been implicated in the pharmacogenetics of ALL therapy were analyzed by TotalPlex amplification and SNP genotyping. We successfully amplified specific gene fragments using 16 pairs of primers in one PCR reaction tube with minimal spurious amplification products using TotalPlex amplification coupled to a multiplexed bead array detection system. The genotypes of 16 loci from 34 different genomic DNA (gDNA) samples derived using the TotalPlex system were consistent with the results of several standard genotyping methods, including automatic sequencing, PCR restriction fragment length polymorphism (RFLP) analysis, PCR, and allele-specific PCR (AS-PCR). Thus, the TotalPlex system represents a useful method of amplification that can improve the time, cost, and sample size required for high-throughput pharmacogenetic analysis of SNPs.

journal_name

Mol Cell Probes

journal_title

Molecular and cellular probes

authors

Kang HJ,Oh Y,Chun SM,Seo YJ,Shin HY,Kim CW,Ahn HS,Han BD

doi

10.1016/j.mcp.2008.02.001

subject

Has Abstract

pub_date

2008-06-01 00:00:00

pages

193-200

issue

3

eissn

0890-8508

issn

1096-1194

pii

S0890-8508(08)00020-0

journal_volume

22

pub_type

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