Sequence alterations can mask each other's presence during screening with SSCP or heteroduplex analysis: BRCA genes as examples

For mutation detection, various screening techniques are widely used becaus e DNA sequencing, the gold-standard method, is still considered to be expen sive and laborious for high-throughput screening. Single-strand conformatio n polymorphism (SSCP) analysis, heteroduplex analysis (HA) and their varian t techniques are popular and frequently used for this purpose. It is widely accepted that when searching for unknown sequence variations, any revealed distinct pattern should always be sequenced. We give examples here of the BRCA1 and BRCA2 genes where the SSCP/HA techniques can produce ambiguous pr edictions if used to detect known genetic variants compared to positive con trols. Using direct DNA sequencing, we provide evidence that in such cases, mutations or polymorphisms can mask each other's presence. This phenomenon can often influence the results of any DNA testing because genetic variati ons such as single-nucleotide polymorphisms occur frequently in the human g enome. We suggest that evert in the case of known electrophoretic patterns of well-characterized genetic alterations, every sequence alteration should be confirmed by direct DNA sequencing, especially if genetic testing is ca rried out for diagnostic purposes.