Chemical methods for the site-specific cleavage of genomic DNA

Abstract

Pyrimidine oligodeoxyribonucleotides bind duplex DNA at homopurine sites to form a triple helix structure. The pyrimidine oligodeoxyribonucleotide is oriented in the major groove of DNA parallel to the Watson-Crick purine strand. Specificity is due to Hoogsteen hydrogen bonds wherein T recognizes A-T base pairs (T•AT triplet) and protonated C recognizes GC base pairs (C + GC triplet). Oligonucleotides 18 bases in length, equipped with a DNA cleaving function EDTA•Fe at the 5' end. cause sequence specific double strand breaks at one site in the 48.5 kbp bacteriophage λ genome. Due to the length of the recognition site, in a formal sense. this is 10^6 times more sequence-specific that restriction enzymes. The triple helix motif can be extended from homopurine target sequences to mixed sequences. Oligonucleotide directed triple helix formation could be useful for mapping genomic DNA.

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