A new thermostable DNA polymerase mixture for efficient amplification of long DNA fragments.
The thermostable DNA polymerases have been used for amplification of DNA fragments since the invention of PCR. The constraint on the maximum size of the amplified fragments can be solved to certain level by the use of unbalanced mixtures of non-proofreading and proofreading thermostable DNA polymerases. In this study, we tested the use of a mixtures of N-terminal deletional variant of Taq polymerase - Klentaq278 and Tne polymerase from Thermotoga neapolitana. Klentaq278 and Tne polymerase genes were cloned and expressed in different expression vectors under tac promoter. The most efficient ratio of Klentaq278/Tne polymerase for amplification was 10 : 1. The polymerase mixture of Klentaq278 and Tne polymerase is very effective in amplification of DNA fragments for up to 8 kb and is useful addition to a DNA polymerases used in long-range PCR.
Davalieva
K G
KG
Research Centre for Genetic Engineering and Biotechnology, Macedonian Academy of Sciences and Arts Krste Misirkov 2, POB 428, 1000 Skopje, Republic of Macedonia.
Efremov
G D
GD
D004247:DNA / Q000096:biosynthesis*; D004256:DNA Polymerase I / Q000737:chemistry* / Q000235:genetics; D017931:DNA Primers; D004795:Enzyme Stability; D006801:Humans; D016133:Polymerase Chain Reaction; D011994:Recombinant Proteins / Q000737:chemistry / Q000235:genetics; D019914:Taq Polymerase / Q000737:chemistry* / Q000235:genetics; D046949:Thermotoga neapolitana / Q000201:enzymology*
D017931:DNA Primers; D011994:Recombinant Proteins; D004247:DNA; D019914:Taq Polymerase; D004256:DNA Polymerase I