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Mutant T7 RNA polymerase is capable of catalyzing DNA primer extension reaction
Author(s) -
Rusakova E.E,
Tunitskaya V.L,
Memelova L.V,
Kochetkova S.V,
Kostyuk D.A,
Kochetkov S.N
Publication year - 1998
Publication title -
febs letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.593
H-Index - 257
eISSN - 1873-3468
pISSN - 0014-5793
DOI - 10.1016/s0014-5793(98)00058-1
Subject(s) - primer extension , primer (cosmetics) , t7 rna polymerase , transcription (linguistics) , dna , chemistry , microbiology and biotechnology , overlap extension polymerase chain reaction , rna , polymerase , nucleotide , mutant , biology , biochemistry , base sequence , gene , escherichia coli , bacteriophage , linguistics , philosophy , organic chemistry , plasmid
The mutant T7 RNA polymerase (T7 RNAP), containing two substitutions (Y639F, S641A) was earlier shown to utilize both rNTP and dNTP in a transcription‐like reaction. In this report the ability of the enzyme to catalyze DNA primer extension reaction was demonstrated. The efficiency of the reaction essentially depended on the type of the primer sequence, and was significantly higher if the primer coincided with the T7 promoter non‐coding sequence. In this case the primer extension reaction proceeded along with de novo RNA synthesis. The length of the product did not exceed 8 nucleotides, indicating that the primer extension reaction proceeds according to the mechanism of the T7 RNAP‐catalyzed abortive transcription.