TRIPLEX-MEDIATED, IN-VITRO TARGETING OF PSORALEN PHOTOADDUCTS WITHIN THE GENOME OF A TRANSGENIC MOUSE

Light-activated psoralens can covalently modify DNA and are widely use d to study nucleic acid secondary structure and mutagenesis. Sequence specificity can be added to the photoaddition reaction by attaching th e psoralen to an oligonucleotide designed to recognize a double-strand ed DNA binding site through formation of a triple helix. We have previ ously used this strategy to study targeted psoralen modification of a tripler binding site within the bacterial supF gene carried in viral g enomes. In the present work we report the targeting of psoralen photoa dducts in vitro to a specific site in the genome of a transgenic mouse . Both 10 base and 16 base oligonucleotide-psoralen conjugates were ca pable of sequence-specific modification of genomic mouse DNA, while a truncated 8 base conjugate was not. Light activation was necessary, an d a dose dependence was demonstrated for target site modification and mutagenesis. The 10 base conjugate rapidly found its target, with sequ ence-specific binding occurring after just 10 min incubation in the pr esence of mouse DNA. The ability to target psoralen photoadducts withi n mammalian genomes may prove useful in the study of chromatin structu re and DNA repair. Moreover, this work may lead to potential in vivo a pplications of targeted psoralen modification.