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

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Authors
Kang, Hyoung Jin; Oh, Yongtaek; Chun, Sung-Min; Seo, Young Jin; Shin, Hee Young; Kim, Chul Woo; Ahn, Hyo Seop; Han, Byoung-Don
Issue Date
2008-02-21
Publisher
Elsevier
Citation
Mol Cell Probes 2008;22:193-200
Keywords
TotalPlex; Bulging primer; Pharmacogenetics; 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.
ISSN
0890-8508
Language
English
URI
http://hdl.handle.net/10371/3708
DOI
https://doi.org/10.1016/j.mcp.2008.02.001
https://doi.org/10.1016/j.mcp.2008.02.001
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College of Medicine/School of Medicine (의과대학/대학원)Pathology (병리학전공)Journal Papers (저널논문_병리학전공)
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