Thermostable DNA Polymerase-encoding Gene from Aquifex pyrophilus and Amino Acid Sequence Deduced Therefrom

摘要

The present invention relates to amino acid sequence of the gene and deduced therefrom of a heat-resistant DNA polymerase (thermostable DNA polymerase) separated from the pillar's (Aquifex pyrophilus) to Aquitania Pecs pie. The present inventors Aquitania Pecs baby duck carcass to thermostable DNA polymerase from the genomic DNA of the filler's in Aquitania Pecs pie using the DNA polymerase gene as probe enzyme of (Aquifex aeolicus)(Aquifex pyrophilus DNA polymerase, Apy DNA polymerase) containing the gene It was selected fragment. Electrical segment Localization of hybridization was cloned into the E. coli colonies hive method Apy DNA polymerase enzyme screening the positive clones containing the gene fragment, and then, by separating the Apy DNA polymerase gene fragments from the clones found to be positive electropositive clones Apy DNA polymerase was confirmed that the fragment containing the gene. Then, were determined the nucleotide sequence of Apy DNA polymerase gene segments, and the deduced amino acid sequence from the electric base sequence, Apy DNA polymerase gene is composed of 1725 bp including the stop codon, encoding a protein composed of 574 amino acids and it was confirmed that.Apy conventional heat-resistant DNA polymerase, the amino acid sequence of a DNA polymerase enzyme and E. coli DNA polymerase amino acid sequence and comparative analysis results of the I, Apy DNA polymerase of the present invention is a high sequence homology to conventional DNA polymerase has been confirmed to be a new heat-resistant DNA polymerase, it was estimated to have also corrected activity (proof-reading activity) that increases the accuracy at the time of DNA polymerase with the DNA polymerase activity (DNA polymerization activity).Apy DNA polymerase gene of the invention is a recombinant thermostable DNA polymerase of more improved high efficiency, high functional features compared to the original enzyme by protein engineering techniques through the industrial mass production, or genetic modification with a heat-resistant DNA polymerase It could be useful in the production.