A simple multiplex PCR method for the concurrent detection of three CYP2C8 variants

• DOI: 10.1016/j.cccn.2004.06.024
• Publication Date: 2004-09-08T14:32:55Z
• Authors: Y.D. Muthiah; W.L. Lee; L.K. Teh; C.E. Ong; M.Z. Salleh; R. Ismail
• Keywords: CYP2C8; genomic DNA; Variants of PCR; Multiplex

Cytochrome P450 (CYP) 2C8 is a principle enzyme responsible for the metabolism of many clinically important drugs as well as endogenous compounds such as arachidonic acid. The enzyme is genetically polymorphic but a simple method is not available to study its genetic polymorphism. We developed and optimized a variant-specific PCR techniques to detect CYP2C8*2, CYP2C8*3 and CYP2C8*4.Genomic DNA was extracted from blood using standard extraction methods. A two-step PCR method was developed to detect simultaneously three CYP2C8 variants. In the first PCR (PCR1), specific regions from exons 3, 5 and 8 of the CYP2C8 gene were amplified. The products were used as templates in parallel alleles-specific PCR (PCR2). This method was tested against DNA samples obtained from 57 healthy Malaysian volunteers.The bands of interest were successfully amplified. This method showed specific and reproducible results when tested on healthy volunteers. DNA sequencing further confirmed genotype results obtained from current method.We have successfully developed and optimized a multiplex PCR method suitable for use in population studies of CYP2C8 polymorphism.