Excision repair of DNA is a major cellular mechanism to correct DNA lesions produced by a variety of physical and chemical agents. The availability of a cell-free system is expected to provide a powerful tool with which to study the complex biochemical pathways of excision repair in eukaryotes. Here we describe such a cell-free system in the yeast Saccharomyces cerevisiae that supports both nucleotide excision repair (NER) and base excision repair (BER) in vitro. Detailed methods and protocols are provided for the preparation of yeast cell-free extracts, damaged DNA substrates, in vitro NER assays, and in vitro BER assays, with an emphasis on technical aspects. NER and BER detected in this cell-free system apparently reflect the complete repair pathways.
ASJC Scopus subject areas
- Molecular Biology
- Biochemistry, Genetics and Molecular Biology(all)