DOE Human Genome Program Contractor-Grantee
25. High-Speed High-Throughput Mutation Detection
Qiufeng Gao, Ho-Ming Pang, and Edward S. Yeung
Ames Laboratory, Iowa State University, Ames, IA 50011
Single-nucleotide polymorphism (SNP) detection has been the focus of much attention recently. Although many methods have been reported, low-cost, high-throughput and high-detection-rate methods are still in demand. We present a fast and reliable mutation detection scheme based on temperature-gradient capillary electrophoresis. A large temperature gradient (10 °C) was applied with a precision of 0.02 °C and a temperature ramp of 0.7 °C/min. Multiple unlabeled samples from PCR reaction were injected and analyzed. Ethidium bromide was used as the intercalating dye for laser-induced fluorescence detection. The mutations were identified by comparing the electrophoretic patterns of the heteroduplex with that of a homoduplex reference without prior knowledge of the DNA sequence. Mutations in all five test samples were successfully detected with high confidence. This scheme is demonstrated in 96-capillary array electrophoresis for screening single-point polymorphism in large numbers of samples.
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