Cloning and targeted mutagenesis of a recA gene required for the extreme DNA damage-resistance of Deinococcus radiodurans

Deinococcus (D.) radiodurans and other members of the same bacterial family are extremely resistant to ionizing radiation, ultraviolet (UV)-radiation and many other DNA-damaging chemical agents. This unusual resistance stems from their extraordinarily efficient DNA repair mechanisms. A mutant strain (strain rec30) generated through chemical mutagenesis has been found to be both DNA-damage sensitive and natural transformation-defficient. Through gene complementation with wild-type (wt) genomic DNA, a gene with extended homology to the Escherichia coli recA was cloned. Transformation of the cloned deinococccal recA gene restored both the DNA-damage resistance and full natural transformation competence of the rec30 mutant. Targeted insertional mutagenesis of the deinococccal recA gene in wt D. radiodurans generated a mutant strain that was phenotypically indistinguishable from strain rec30. These results indicate that the observed phenotypic changes in strain rec30 were caused by a defect in its recA gene, and a RecA-mediated recombination repair pathway plays a critical role in the DNA repair of D. radiodurans.