Sequence-specific targeting and covalent modification of human genomic DNA

We compare two techniques which enable selective, nucleotide-specific covalent modification of human genomic DNA, as assayed by quantitative ligation-mediated PCR. In the first, a purine motif tripler-forming oligonucleotide with a terminally appended chlorambucil was shown to label a target guanine residue adjacent to its binding site in 80% efficiency at 0.5 mu M. Efficiency was higher in the presence of the triplex-stabilizing intercalator coralyne, In the second method, an oligonucleotide targeting a site containing all four bases and bearing chlorambucil on an interior base was shown to efficiently react with a specific nucleotide in the target sequence, The targeted sequence in these cases was in the DQ beta 1*0302 allele of the MHC II locus.