engleski

The accuracy of ACE (angiotensin-converting enzyme) insertion/deletion polymorphism genotyping

Rationale: The insertion/deletion polymorphism (I/D) in the intron 16 of the angiotensin- converting enzyme (ACE) is associated with plasma ACE activity and may confer an increased risk to hypertension, CHD and stroke. Genotyping error for ACE I/D polymorphism has been widely recognized and is estimated to account for almost 40% of misclassification. The most probable cause for this error is preferential amplification of D allele over I allele. To compensate this error, several modifications have so far been suggested. The aim of this study was to assess the comparability of three modified ACE I/D genotyping protocols with original genotyping method reported by Rigat. Materials and Methods: 200 samples were genotyped for ACE I/D polymorphism, using the original PCR protocol reported by Rigat et al. slightly modified for primer sequence and cycling conditions. Three additional modifications were used: 1) step-down PCR protocol (amplification temperature gradually decreasing) ; 2) DMSO protocol (4% dimethyl sulfoxide was added to reaction mixture) and 3) protocol with insertion specific primers. I/I, I/D and D/D control samples were used per every 20 patient samples. PCR fragments were separated electrophoretically. Results: Genotype frequencies using the original method were as follows: 19.5% I/I, 54.0% I/D and 28.5% D/D. We were not able to confirm the genotyping error of the original method by Rigat. The concordance rate of original method with all three modified methods was 100%. Conclusion: If properly optimized, original method by Rigat does not suffer from genotyping error.

angiotensin-converting enzyme; polymorphism; genotyping error

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