Sequence-Specific Double-Strand Alkylation and Cleavage of DNA Mediated by Triple-Helix Formation
Attachment of the nondiffusible electrophile N-bromoacetyl to the 5-position of a thymine at the 5′-end of a pyrimidine oligodeoxyribonucleotide affords sequence specific alkylation of a guanine two base pairs to the 5′-side of a local triple-helix complex in >96% yield. N-Bromoacetyloligodeoxyribonucleotides bind adjacent inverted purine tracts on double-helical DNA by triple-helix formation and alkylate single guanine positions on opposite strands at 37 °C (pH 7.4). After depurination, double-strand cleavage at a single site within plasmid DNA (4 kp in size) occurs in greater than 85% yield. The resulting DNA fragments from site-specific alkylation and cleavage can be ligated with DNA fragments generated by restriction endonuclease digestion. This nonenzymatic approach which couples sequence-specific recognition with sequence-dependent cleavage affords double-strand site-specific cleavage in megabase size DNA. A yeast chromosome, 340000 base pairs in size, was cleaved at a single site in 85-90% yield. © 1992, American Chemical Society. All rights reserved.
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Published In
DOI
EISSN
ISSN
Publication Date
Volume
Issue
Start / End Page
Related Subject Headings
- General Chemistry
- 40 Engineering
- 34 Chemical sciences
- 03 Chemical Sciences