Analysis of the Fine Structure of the Repair of Anti−benzo[a]pyrene-7,8-diol-9,10-epoxide-DNA Adducts in Mammalian Cells by Laser-Induced Strand Cleavage

Abstract The repair of DNA adducts formed by reaction of anti−benzo[a]pyrene-7,8-diol-9,10-epoxide (7-hydroxyl and epoxide oxygen trans) [BaPDE] with DNA in the multiple copy dihydrofolate reductase (DHER) gene of the B11 Chinese hamster ovary cell line was investigated by restriction enzyme digestion, alkaline agarose gel electrophoresis, and probing for the presence of specific full length restriction fragments with 32P-riboprobes. Cleavage of fragments that contained BaPDE-DNA adducts was carried out photochemically with 355 nm light from a Nd: YAG laser. Exposure of KpnI restricted DNA modified to various extents with BaPDE to a laser intensity of 40 mW for 5 to 10 minutes resulted in quantitative cleavage of fragments containing BaPDE adducts, with minimal cleavage of unmodified fragments. In cells treated with 2.5 μM BaPDE very little repair was observed in a 14 kb fragment at the 5' end of the DHFR gene over a 24 hour period for both the transcribed and nontranscribed strands. This photochemical cl...

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