Alkylation of individual genes in rat liver by the carcinogen N-nitrosodimethylamine.
A method for quantifying alkylated bases in defined gene sequences of rat liver following carcinogen treatment is described. After restriction, the genomic DNA is heated to 65 degrees at pH 8.0 to cause depurination of 7-alkylguanine and 3-alkyladenine residues. Reaction with spermidine then generates strand breaks. After gel electrophoresis and Southern transfer, sequences of interest are visualized using specific probes. The presence of strand breaks in restriction fragments reduces their intensity compared to unmethylated fragments. The method was applied to show that the transcriptionally active albumin gene is damaged by the hepatocarcinogen N-nitrosodimethylamine to a much greater extent than the untranscribed IgE gene in rat liver.
Department of Medical Biophysics, University of Toronto, Canada.
J R
JR
Milligan
M C
MC
Archer
D000418:Albumins / Q000235:genetics*; D000478:Alkylation; D000818:Animals; D004249:DNA Damage; D004128:Dimethylnitrosamine; D005786:Gene Expression Regulation / Q000187:drug effects; D007073:Immunoglobulin E / Q000235:genetics*; D008099:Liver / Q000378:metabolism*; D008297:Male; D051381:Rats; D011919:Rats, Inbred Strains
D000418:Albumins; D007073:Immunoglobulin E; D004128:Dimethylnitrosamine