Measurement of DNA damage and repair in the lambda LIZ transgene in a Big Blue rat cell line by quantitative PCR.
DNA damage and its subsequent repair occur heterogeneously throughout the genome, which reflects the nature of the damaging agents, gross chromosomal structure, the specific nucleotide sequence, and transcriptional status. We selected to investigate the repair of DNA damage in an artificial, transgenic situation. Here we report the repair of UV and BPDE-induced DNA damage in the nontranscribed lambda construct of the Big Blue rat-2 transgenic cell line. This was determined by quantitative PCR (QPCR) using genomic DNA isolated at specific times following treatment with UV or BPDE. The results indicate that, despite the absence of transcription, lesions induced in the lacI-containing lambda insert by UV and BPDE are efficiently repaired. The half-life of the polymerase-blocking lesions is 4.2 and 5.5 hr for UV and BPDE induced lesions, respectively. This is an important observation vis-a-vis the use of this transgene as a model for studies of mutational mechanism.
Centre for Environmental Health, University of Victoria, British Columbia, Canada.
H
H
Yang
G P
GP
Kotturi
J G
JG
de Boer
B W
BW
Glickman
D015123:7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide / Q000378:metabolism; D000818:Animals; D002460:Cell Line; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics / Q000528:radiation effects; D018736:DNA Adducts / Q000378:metabolism; D004249:DNA Damage; D004260:DNA Repair; D004622:Embryo, Mammalian; D005347:Fibroblasts / Q000187:drug effects / Q000378:metabolism / Q000528:radiation effects; D007700:Kinetics; D016133:Polymerase Chain Reaction / Q000379:methods*; D051381:Rats; D019914:Taq Polymerase / Q000737:chemistry; D019076:Transgenes / Q000187:drug effects / Q000235:genetics* / Q000528:radiation effects; D014466:Ultraviolet Rays
D018736:DNA Adducts; D015123:7,8-Dihydro-7,8-dihydroxybenzo(a)pyrene 9,10-oxide; D019914:Taq Polymerase