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Published on: April 1, 2019
A simple method for the detection of CYP2C9 polymorphisms: nested allele-specific multiplex polymerase chain reaction
Z Zainuddin1, L K Teh, A W M Suhaimi
1Pharmacogenetics Study Group, Department of Pharmacology, School of Medical Sciences, Universiti Sains Malaysia, 16150 Kubang Kerian, Kelantan, Malaysia.
Background:
Cytochrome P4502C9 (CYP2C9), a principle drug-metabolizing enzyme is polymorphic in humans and is responsible for important pharmacokinetic and pharmacodynamic variations of CYP2C9 substrates. We developed an allele-specific multiplex polymerase chain reaction (PCR) method for the detection of common CYP2C9 alleles.
Method:
Genomic DNA was extracted from blood obtained from 40 unrelated healthy Malaysian Indian volunteers. The DNA was subjected to a first PCR that was used to amplify both exons 3 and 7 simultaneously in one reaction tube and a second PCR that was used to detect the polymorphic sites of CYP2C9 alleles using allele-specific primers. Sequencing was performed to validate the test results.
Results:
We were successful in amplifying the fragments of interest from the DNA samples. The method was also reproducible and specific. The amplified sequences showed 100% homology to CYP2C9 sequence.
Conclusion:
This is the first nested allele-specific multiplex PCR method reported to allow for the simultaneously detection of five CYP2C9 alleles.
