Cloning a eukaryotic DNA glycosylase repair gene by the suppression of a DNA repair defect in Escherichia coli.
If eukaryotic genes could protect bacteria with defects in DNA repair, this effect could be exploited for the isolation of eukaryotic DNA repair genes. We have thus cloned a DNA repair gene from Saccharomyces cerevisiae that directs the synthesis of a DNA glycosylase that specifically releases 3-methyladenine from alkylated DNA and in so doing protects alkylation-sensitive Escherichia coli from killing by methylating agents. The cloned yeast gene was then used to generate a mutant strain of S. cerevisiae that carries a defect in the glycosylase gene and is extremely sensitive to DNA methylation. This approach may allow the isolation of a large number of eukaryotic DNA repair genes.
Chen
J
J
Harvard School of Public Health, Charles A. Dana Laboratory of Toxicology, Boston, MA 02115.
Derfler
B
B
Maskati
A
A
Samson
L
L
D015152:Blotting, Northern; D015139:Blotting, Southern; D003001:Cloning, Molecular / Q000379:methods; D004249:DNA Damage; D045647:DNA Glycosylases; D004260:DNA Repair; D004351:Drug Resistance; D004926:Escherichia coli / Q000201:enzymology / Q000235:genetics* / Q000254:growth & development; D005800:Genes, Fungal; D008741:Methyl Methanesulfonate / Q000494:pharmacology; D009699:N-Glycosyl Hydrolases / Q000235:genetics* / Q000378:metabolism; D011994:Recombinant Proteins / Q000378:metabolism; D015183:Restriction Mapping; D012441:Saccharomyces cerevisiae / Q000187:drug effects / Q000201:enzymology / Q000235:genetics*
D011994:Recombinant Proteins; D008741:Methyl Methanesulfonate; C021565:3-methyladenine-DNA glycosylase; D045647:DNA Glycosylases; D009699:N-Glycosyl Hydrolases