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CLEAVAGE OF PHOSPHOROTHIOATE-CONTAINING OLIGONUCLEOTIDES AND DNA BY RESTRICTION ENDONUCLEASES
Eckstein, F. Conolly, B. A. Potter, B. V. L.
Eckstein, F.
Conolly, B. A.
Potter, B. V. L.
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Issue Date
1987
Submitted date
2023-04-19
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Abstract
The oligonucleotide d(GGsAATTCC) containing the recognition sequence of the Eco R1 restriction endonuclease with a phosphorothioate group at the site of cleavage was synthesized by two approaches both based on the polymer support phosphoroamidite method. Only the Rp-diastereomer was cleaved by Eco R1, at a rate approximately 20 times slower than the all-phosphate-containing octamer. To study the interaction of restriction endonucleases with phosphorothioate internucleotidic linkages double stranded Mi3mp2 DNA was prepared in which the (+)strand contained only phosphate groups but the (-)strand phosphate groups as well as base specifically introduced phosphorothioate groups of the Rp-configuration. This hybrid DNA was cleaved by several restriction enzymes. The results obtained allow the classification of these enzymes into three groups: those which produce nicked DNA as an isolatable intermediate, those where the nicked DNA is the final product and a third where only linearised DNA can be detected. Particularly the second class of enzymes should be of interest for the manipulation of DNA.
Citation
Chemical synthesis in molecular biology, 23 ff
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Book chapter
conference paper
conference paper
Language
en
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GBF Monographs, Vol. 8
ISSN
0930-4320
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Attribution-NonCommercial-ShareAlike 4.0 International