Expression of Escherichia Coli Uvr Genes in Mammalian Cells. Progress Report

摘要

The E. coli uvrA, uvrB and uvrC gene products are required to carry out the incision step in excision repair of damaged DNA. The range of sensitivity to DNA damage in E. coli uvrA uvrB, and uvrC mutants is similar to that of cells derived from patients with xeroderma pigmentosum (XP). Our goal is to introduce the E. coli uvrA, uvrB, and uvrC genes into SV40-transformed XP cells and to examine whether these genes are able to genetically complement the repair deficiency in XP cells. Ecogpt was used as a selection marker for uvr gene transfection into XP cells. The uvr genes were cloned into composite pBR322-SV40 and Ecogpt vectors in which each E. coli gene is flanked by individual SV40 regulatory elements. We have also constructed vectors in which the uvr gene and the gpt are in tandem and flanked by one set of SV40 regulatory elements. SV40-transformed XP cells of complementation group A (XP12BE) were transfected with pSV2uvrASV2gpt, a vector of the former configuration. Southern gel analysis of the resulting gpt exp + cells has revealed one colony in which the uvrA gene with its SV40 elements has been integrated into the chromosomal DNA intact. (ERA citation 08:052390)