We have developed methylation, sensitive, single-strand conformation analysis (MS-SSCA) as a method of screening for methylation changes. Bisulfite modification converts cytosines to thymines, but methylated cytosines remain unchanged. This modification creates sequence differences between methylated and unmethylated samples, which can be resolved by SSCA. SSCA is 70-95% efficient at detecting single base changes in a fragment. As bisulfite modification of methylated DNA would typically involve several base changes in a fragment, the efficiency of detecting methylation using MS-SSCA could approach 100%. We applied this method to analyze the BRCA1 promoter CpG island in breast cancer samples. About 20% of sporadic breast cancers are hypermethylated at the BRCA1 promoter CpG island. MS-SSCA rapidly detected those tumors that had previously been shown to be methylated by Southern blotting. The variant bands detected by SSCA were analyzed by sequencing and shown to be methylated. MS-SSCA is a simple method for screening large numbers of samples for methylation and can accelerate genomic sequencing, as all bands can be isolated and sequenced directly.
|Number of pages||5|
|Publication status||Published - Oct 1999|
- Single-strand conformation analysis