Cleavage of functionalized DNA containing 5-modified pyrimidines by type II restriction endonucleases
Language English Country Germany Media print-electronic
Document type Journal Article, Research Support, Non-U.S. Gov't
- MeSH
- DNA metabolism MeSH
- Pyrimidines chemistry MeSH
- Deoxyribonucleases, Type II Site-Specific metabolism MeSH
- DNA Restriction Enzymes metabolism MeSH
- DNA Cleavage MeSH
- Publication type
- Journal Article MeSH
- Research Support, Non-U.S. Gov't MeSH
- Names of Substances
- DNA MeSH
- Pyrimidines MeSH
- Deoxyribonucleases, Type II Site-Specific MeSH
- DNA Restriction Enzymes MeSH
A series of six pyrimidine-modified dNTPs--5-ethynyl-, 5-phenyl-, and 5-(3-nitrophenyl)deoxycitidine and -deoxyuridine triphosphates--were prepared and incorporated by primer extension with Vent (exo-)polymerase to specific DNA sequences within or next to the recognition sequences of selected restriction endonucleases. The cleavage of these pyrimidine-modified DNA sequences by 13 restriction enzymes was then studied. Whereas the presence of any modified C within the target sequence completely prevented any restriction cleavage, most enzymes tolerated the presence of 5-ethynylU and two of them even the presence of 5-phenyl- and 5-(3-nitrophenyl)U. Modifications outside the recognition sequence were tolerated except in the case of phenyl derivatives with the PvuII enzyme. 5-EthynylC was used for protection of the recognition sequence from cleavage in the presence of the second unmodified copy of the same sequence that was cleaved.
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