Nucleic Acids Research, 1984, Vol. 12, No. 24 9237-9248
© 1984
MOLECULAR BIOLOGY |
The complete pattern of mutagenesis arising from the repair of site-specific psoralen crosslinks: analysis by oligonucleotide hybridizatlon
Department of Human Genetics and Development, College of Physicians and Surgeons, Columbia University New York, NY 10032, USA
Received November 14, 1984. Accepted November 23, 1984.
Psoralen crosslinks were site-specifically placed in plasmid pBR322 near the BamHI site in the tet gene by enzymatically inserting mercurated nucleotides and reacting at the target site with a sulfhydryl-containing psoralen. The damaged plasmid was repaired in SOS-induced E. coli cells. Mutants were detected by colony hybridization to oligonucleotides in the target region, and their sequences were determined. The mutations are all base substitutions, 80% transitions and 20% transversions, similar to the mutations previously identified by the loss of tetracycline resistance. However, the mutation sites detected by a physical method, unconstrained by phenotypic changes, follow a broader distribution than those identified genetically. They occur primarily at favored psoralen crosslinking sites, where T-T and T-C interstrand crosslinks can be formed. A majority of these mutations are silent.