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 triplex-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 microM. 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 DQbeta1*0302 allele of the MHC II locus.
Belousov
E S
ES
Epoch Pharmaceuticals, Inc., 1725 220th Street SE, #104, Bothell, WA 98021, USA.
Afonina
I A
IA
Podyminogin
M A
MA
Gamper
H B
HB
Reed
M W
MW
Wydro
R M
RM
Meyer
R B
RB
D000483:Alleles; D001483:Base Sequence; D001665:Binding Sites; D002699:Chlorambucil; D004247:DNA / Q000737:chemistry*; D018390:Gene Targeting; D005802:Genes, MHC Class II; D006683:HLA-DQ Antigens / Q000235:genetics; D059866:HLA-DQ beta-Chains; D006801:Humans; D008969:Molecular Sequence Data; D009838:Oligodeoxyribonucleotides / Q000737:chemistry; D016133:Polymerase Chain Reaction; D011687:Purines / Q000737:chemistry
D006683:HLA-DQ Antigens; D059866:HLA-DQ beta-Chains; C069051:HLA-DQB1 antigen; D009838:Oligodeoxyribonucleotides; D011687:Purines; C051802:triplex DNA; D002699:Chlorambucil; D004247:DNA