Open AccessAllele‐specific polymerase chain reaction for genotyping human cytochrome P450 2E1
Author(s) -
Sohda Tetsuro
Publication year - 1999
Publication title -
journal of clinical laboratory analysis
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.536
H-Index - 50
eISSN - 1098-2825
pISSN - 0887-8013
DOI - 10.1002/(sici)1098-2825(1999)13:5<205::aid-jcla2>3.0.co;2-m
Subject(s) - genotyping , polymerase chain reaction , microbiology and biotechnology , genotype , multiplex polymerase chain reaction , biology , variants of pcr , allele , restriction enzyme , hot start pcr , restriction fragment length polymorphism , primer dimer , genetics , oligonucleotide , polymerase chain reaction optimization , gene
Allele‐specific polymerase chain reaction (AS‐PCR) was applied to investigate the cytochrome P450 2E1 (CYP2E1) genotype. AS‐PCR is a competitive multiplex PCR method in which PCR amplification is successfully performed only by using the sequence of 3′ oligonucleotide ends as a DNA template in order to obtain an absolutely complementary product. I was able to produce allele‐specific primers whose 3′ ends had the base specific to Pst I polymorphism located within the 5′‐flanking region of the CYP2E1 gene. Electrophoresis of the products showed that bands derived from common PCR products, allele C1 and C2, were clearly separate from each other due to the difference in the size of the products. I tested 102 unrelated Japanese individuals, and the results of both restriction fragment length polymorphism (RFLP) by Pst I or Rsa I and direct sequencing were in complete agreement with those of AS‐PCR. These results lead me to conclude that AS‐PCR is a simple and useful technique for investigating CYP2E1 genotype. J. Clin. Lab. Anal. 13:205–208, 1999. © 1999 Wiley‐Liss, Inc.