Amplification of real-time high resolution melting analysis PCR method for polycystic kidney disease (PKD) gene mutations in autosomal dominant polycystic kidney disease patients

Abstract

PKD1 and PKD2 are the two genes responsible for the development of autosomal dominant polycystic kidney disease (ADPKD). PKD1 gene mutations accounts for ≈85% of all ADPKD cases, while the remaining ≈15% of cases is caused by mutations in the PKD2 gene. Genotyping for PKD1 and PKD2 mutations was usually identified using conventional polymerase chain reaction (PCR) or PCR-single stranded conformation polymorphisms (SSCP) methods. In this study, we assessed the usefulness of eight common primers amplifying the respective genes in real-time high resolution melting analysis PCR (real-time HRMA PCR) in terms of time, cost and sensitivity with respect to PCR-SSCP method. We found that case sample can easily be differentiated from control sample by melting curve profile difference, although a primer was found to be less useful. We concluded that real-time HRMA PCR is a rapid and sensitive method to categorize samples based on the melting curve profiles with comparable sensitivity to conventional PCR-SSCP.

Key words: Autosomal dominant polycystic kidney disease (ADPKD), real-time PCR, high resolution melting analysis, PKD1, PKD2.