title: Construction of a functional cDNA clone of the hamster ERCC2 DNA repair and transcription gene.
abstract: The complete hamster ERCC2 cDNA was constructed in a plasmid vector from clones of three overlapping reverse transcribed/polymerase chain reaction amplified fragments using unique restriction enzyme recognition sites within the regions of overlap. This complete cDNA insert was then cloned into a mammalian expression vector, pcD2E, and tested for function by the ability to confer UV resistance to the ERCC2 mutant CHO cell line UV5. Site-specific mutagenesis was used to introduce the G347-->A and G1844-->A changes resulting in the Cys116-->Tyr and Gly615-->Glu mutations previously identified in UV5 and UVL-13 (also an ERCC2 mutant CHO cell line), respectively. The 116Tyr and 615Glu plasmids each failed to confer UV resistance to UV5 or UVL-13 cells, respectively, demonstrating that the changes identified are indeed the causative mutations in UV5 and UVL-13.
authors.affiliation: Biology and Biotechnology Research Program, Lawrence Livermore National Laboratory, Livermore, California 94551, USA.
authors.forename: S
authors.initials: S
authors.lastname: Kadkhodayan
authors.forename: E P
authors.initials: EP
authors.lastname: Salazar
authors.forename: J E
authors.initials: JE
authors.lastname: Lamerdin
authors.forename: C A
authors.initials: CA
authors.lastname: Weber
mesh_terms: D000818:Animals; D016466:CHO Cells; D006224:Cricetinae; D004265:DNA Helicases; D004260:DNA Repair / Q000235:genetics*; D018076:DNA, Complementary / Q000235:genetics*; D004268:DNA-Binding Proteins; D005822:Genetic Vectors; D010957:Plasmids; D011506:Proteins / Q000235:genetics*; D014157:Transcription Factors; D014158:Transcription, Genetic; D051759:Xeroderma Pigmentosum Group D Protein
chemical_list: D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D011506:Proteins; D014157:Transcription Factors; D004265:DNA Helicases; D051759:Xeroderma Pigmentosum Group D Protein