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Two-step "hot" PCR amplification of GC-rich avian c-myc sequences
M Schuchard1, G Sarkar, T Ruesink
1Dept. of Biochemistry and Molecular Biology, Mayo Foundation, Rochester, MN 55905.
Biotechniques
|March 1, 1993
Abstract:
A new two-step cycle PCR method has been developed for amplification of GC-rich DNA sequences. Using this method, termed "hot PCR," 111 and 179 bp regions (GC contents of 74% and 76%, respectively) of the avian c-myc proto-oncogene were specifically amplified from cloned and genomic DNA. This method uses high-melting primers (Tm between 70 degrees and 74 degrees C), a two-step cycle that employs a 94 degrees C denaturation step and an annealing-elongation step between 70 degrees and 80 degrees C with or without formamide.