Rapid screening of MMACHC gene mutations by high-resolution melting curve analysis.

Abstract

BackgroundCobalamin (cbl) C is a treatable rare hereditary disorder of cbl metabolism with autosomal recessive inheritance. It is the most common organic acidemia, manifested as methylmalonic academia combined with homocysteinemia. Early screening and diagnosis are important. The mutation spectrum of the MMACHC gene causing cblC varies among populations. The mutation spectrum in Chinese population is notably different from that in other populations.MethodsA PCR followed by high‐resolution melting curve analysis (PCR‐HRM) method covering all coding exons of MMACHC gene was designed to verify 14 pathogenic MMACHC gene variants found in patients with cblC, including all common mutations in Chinese patients with cblC.ResultBy PCR‐HRM analysis, 14 pathogenic variants of MMACHC showed distinctly different melting curves, which were consistent with Sanger sequencing. The homozygous type of the most common mutation c.609G > A (p.Trp203Ter) can also be analyzed by specially designed PCR‐HRM.ConclusionThe established PCR‐HRM method for screening common pathogenic MMACHC variants in Chinese patients with cblC has the advantages of high accuracy, high throughput, low cost, and high speed. It is suitable for the large‐sample screening of suspected children with methylmalonic acidemia and carriers in population.