Detection of TPMT*3C gene polymorphism by polymerase chain reaction (PCR) in related to azathioprine medication

Azathioprine is an immunosuppressive drug used to treat inflammatory bowel diseases, rheumatoid arthritis and to prevent rejection in organ transplantation. It is metabolized mainly into inactive 6-methylmercaptopurine (6-MMP) via thiopurine S-methyltransfersase (TPMT) enzyme. The deficiency of TPMT enzyme due to genetic polymorphism has a higher risk of developing severe, life-threatening myelotoxicity when given standard doses of azathioprine. The Nested PCR method was used to detect the SNPs in TPMT*3C (rsl 142345). It is one of the major allele that causes the activity of TPMT enzyme to be low. The study focused on the optimization of the Nested PCR where the parameters were adjusted to obtain the optimized condition. The parameters altered in this study include the concentration of DNA template, annealing temperature and the volume of primers. The amplified PCR product was evaluated by gel electrophoresis and observed under UV light. In conclusion, the optimization of Nested PCR method was not yet achieved hence the mutant and wild type cannot be differentiated. The other parameters including the concentration of MgCh, the number of cycles in PCR execution and concentration of enzyme should be adjusted as well.