Abstract
Saccharomyces cerevisiae (S. cerevisiae) (commonly known as baker’s yeast) is a model organism that has a similar upstream base excision repair (BER) pathway for the repair of methylated bases as that in mammalian cells, and it is very easy to maintain in the laboratory environment. Here, we described a method to prepare cell extracts from yeast to investigate their enzymatic activities. This protocol is a quick and efficient way to make yeast cell extracts without using commercial kits.
Keywords: DNA damage, DNA polymerase, DNA repair, Base excision repair, Ligation failure
Materials and Reagents
Equipment
Procedure
Representative data
The result image that shows dRP lyase, FEN1 excision and DNA deadenylation enzymatic activities in yeast extracts was published as Supplementary Figure 5 in Caglayan et al. (2014).
Notes
Enzymatic activity in yeast cell extracts may be lost after continuous vortexing if samples warm up. It might be necessary to extend the alternating cooling steps depending on the sample.
Recipes
Acknowledgments
This work was supported by the Intramural Research Program of the US National Institutes of Health, National Institute of Environmental Health Sciences (grants Z01 ES050158 and ES050159).
References
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