We have developed a highly versatile platform that performs temperature gradient capillary electrophoresis (TGCE) for mutation/single-nucleotide polymorphism (SNP) detection, sequencing and mutation/SNP genotyping for identification of sequence variants on an automated 24-, 96- or 192-capillary array instrument. In the first mode, multiple DNA samples consisting of homoduplexes and heteroduplexes are separated by CE, during which a temperature gradient is applied that covers all possible temperatures of 50% melting equilibrium (Tms) for the samples. The differences in Tms result in separation of homoduplexes from heteroduplexes, thereby identifying the presence of DNA variants. The sequencing mode is then used to determine the exact location of the mutation/SNPs in the DNA variants. The first two modes allow the rapid identification of variants from the screening of a large number of samples. Only the variants need to be sequenced. The third mode utilizes multiplexed single-base extensions (SBEs) to survey mutations and SNPs at the known sites of DNA sequence. The TGCE approach combined with sequencing and SBE is fast and cost-effective for high-throughput mutation/SNP detection.
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