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African Journal of Biotechnology

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

RM Jas, R Vasudevan, P Ismail, AHA Gafor, S Moin, SA Eshkoor

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.




http://dx.doi.org/10.5897/AJB11.1427
AJOL African Journals Online