Rapid screening of a hotspot variant c.609G>A in MMACHC gene by using PCR-high-resolution melting curve analysis.

Author: Shuxiang LIN ¹ ; Chao WANG ; Xinjie ZHANG ; Xiaowei XU ; Qianqian ZOU ; Chunquan CAI ; Yuqin ZHANG ; Jianbo SHU
Author Information

1. Tianjin Pediatric Research Institute, Tianjin Children's Hospital, Tianjin 300134, China. shjb1981@sina.com.
Publication Type:Journal Article
From: Chinese Journal of Medical Genetics 2020;37(7):759-763
CountryChina
Language:Chinese
Abstract: OBJECTIVE:To carry out genetic testing for two families affected with cobalamin C (cblC) and establish a rapid method for the detection of a hotspot pathogenic variant c.609G>A of the MMACHC gene by using a PCR-high-resolution melting curve (PCR-HRM) method.
METHODS:Genomic DNA was extracted from peripheral blood samples of the probands and their parents. Potential variants of the MMACHC gene was analyzed by Sanger sequencing. The c.609G>A variant of the MMACHC gene was screened among 100 healthy children with the PCR-HRM method.
RESULTS:Sanger sequencing revealed that proband 1 carried compound heterozygous variants c.394C>T and c.609G>A of the MMACHC gene, while proband 2 carried compound heterozygous variants c.482G>A and c.609G>A of the same gene. PCR-HRM analysis of the two probands and the 100 healthy children were consistent with the Sanger sequencing.
CONCLUSION:c.609G>A is a hotspot pathogenic variant of the MMACHC gene. The diagnosis of cblC may be rapidly attained through detection by PCR-HRM.