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PCR genotyping of CYP2D6 gene

Cytochrome P450 (CYP) family presents the most important oxidative enzyme system involved in the metabolism of numerous widely used drugs including antiarrhytmics, antidepressants, beta-blockers, and neuroleptics. More than thirty CYP isoenzymes have been characterized in humans, each with distinct catalytic specificity and unique regulation. The genetic polymorphism has been linked to three classes of phenotypes relating to drug metabolism; extensive (EM), poor (PM) and ultraextensive (UEM) metabolizers resulting in normal, high and low blood levels of the therapeutics. PCR method presents screening method for genetic mutations associated with altered metabolism of drugs by amplification of specific region of the gene of interest followed by digestion of the amplified DNA product with restriction endonucleases and comparison the size of digestion products generated from a DNA substrate amplified from control subject DNA vs. study subject DNAs. Differences in the size of DNA fragments generated as a result of endonuclease digestion, commonly referred as a restriction fragment length polymorphism (RFLP), can be easily evaluated by agarose gel electrophoresis with ethidium bromide staining and UV transillumination. The protocol for the two mutations of the most common allele (CYP2D6A-2,7% and CYP2D6B - 28,6%) of CYP2D6 gene has been described.

cytochrome P450; isoenzymes; CYP2D6; drug metabolism; EM; PM; UEM genotyping; phenotyping; PCR-RFLP

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