Single-stranded nucleic acids differing by a single base form different secondary structures that migrate with different mobilities in a polyacrylamide nondena-turing gel. Orita applied this principle in SSCP analysis to detect sequence differences within the same region of DNA from different individuals.58 Differences in the electrophoretic patterns obtained are determined visually by nonradioac-tive means. Under optimal conditions and with the fragment size of the DNA test sample within the range of 150-200 base pairs, 80-90% of base exchanges are detectable.59
Recently, high-SSCP analysis has been automated for use with capillary electrophoresis.60,61 This method has been used to detect point mutations associated with inherited cardiac disorders including the long QT syndrome and hypertrophic cardiomyopathy. The sensitivity was 100% when 34 different point mutations were analyzed on PCR fragments of 166-1223 base pairs in length. Limited experience indicates that the automated technique has advantages of rapidity, robustness, high resolution, and good reproducibility.
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