High‐sensitivity Epimutations Testing with MS‐HRM

Epimutation is defined as any heritable change in gene activity that does not affect the actual sequence of DNA. One of the most researched epimutations is promoter methylation of breast cancer susceptibility gene BRCA1. This epimutation may arise de novo in somatic tissues, such as breast or ovarian cancer tissue, but has also been shown to be widely distributed throughout tissues. BRCA1 methylation detectable in peripheral blood has been associated with the risk of both breast and ovarian cancer in patients without a germline pathogenic variant status of this gene. The strength of this association suggests that diagnostic screening for the BRCA1 epimutation should be considered with potential implications in cancer risk assessment. However, the detection of BRCA1 epimutation may be challenging, as it is generally detectable at a very low level in blood. Methylation-sensitive high-resolution melting (MS-HRM) was shown to provide a high sensitivity of methylation detection, and as it is a PCR-based method, epimutation screening based on MS-HRM is labor- and cost-effective. Here we describe the MS-HRM protocol for BRCA1 epimutation screening. © 2025 Wiley Periodicals LLC. Basic Protocol: High sensitivity constitutional epimutations testing with MS-HRM.