Allele-specific polymerase chain reaction for genotyping human cytochrome P450 2E1

✍ Scribed by Tetsuro Sohda

• Publisher: John Wiley and Sons
• Year: 1999
• Tongue: English
• Weight: 91 KB
• Volume: 13
• Category: Article
• ISSN: 0887-8013
• DOI: 10.1002/(sici)1098-2825(1999)13:5<205::aid-jcla2>3.0.co;2-m

✦ Synopsis

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 prod-ucts 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.