Abstract
We have developed a rapid method to synthesize radioactively labeled single-stranded DNA probes suitable for strand-specific analysis of single copy genes on Southern blot. Linear PCR with 10 μCi α32P-aATP (3000 Ci/mmol) as the only dATP source enabled us to generate strand-specific DNA probes with high specific activity. The probes synthesized by this method have higher specific activities and the same strand specificity compared to the end-labeled single-stranded DNA probes obtained from single-stranded M13mp18/19 vectors. Application of the method for strand-specific analysis of ultraviolet-induced DNA lesions in defined DNA sequences significantly improved the hybridization signal.
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