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Incorporation of 2′‐Deoxy‐2′‐isonucleoside 5′‐Triphosphates (iNTPs) into DNA by A‐ and B‐Family DNA Polymerases with Different Recognition Mechanisms
Author(s) -
Ogino Takaaki,
Sato Kousuke,
Matsuda Akira
Publication year - 2010
Publication title -
chembiochem
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.05
H-Index - 126
eISSN - 1439-7633
pISSN - 1439-4227
DOI - 10.1002/cbic.201000449
Subject(s) - klenow fragment , primer (cosmetics) , dna polymerase , oligonucleotide , base pair , dna , polymerase , dna polymerase i , nucleotide , nucleobase , chemistry , nucleoside , primase , stereochemistry , biochemistry , biology , microbiology and biotechnology , reverse transcriptase , polymerase chain reaction , gene , organic chemistry , exonuclease
Recently, α‐ L ‐threofuranosyl nucleoside 3′‐triphosphates (tNTPs) have been reported to be incorporated into DNA by DNA polymerases. Isonucleosides especially the 2′‐deoxy‐2′‐isonucleosides, would be considered regioisomers of α‐ L ‐threofuranosyl nucleosides. Therefore, we investigated the synthesis of 2′‐deoxy‐2′‐isonucleoside 5′‐triphosphates (iNTPs) having the four natural nucleobases and their incorporation into primer–template duplexes consisting of oligonucleotides containing natural 2′‐deoxyribonucleosides and 2′‐deoxy‐2′‐isonucleosides by using primer‐extension reactions. We found that Klenow fragment ( exo ‐; an A‐family DNA polymerase) has strict recognition of the shape of nucleoside 5′‐triphosphates and Therminator (a B‐family DNA polymerase) has strict recognition of the shape of primer–template complexes, especially two base pairs upstream of the primer 3′ terminus.