Cloning and purification of large fragment of DNA polymerase I from Geobacillus stearothermophilus and it’s application in isothermal DNA amplification

2017 ◽  
Author(s):  
P. Li ◽  
◽  
A. Amenov ◽  
R. Kalendar ◽  
S. Abeldenov ◽  
...  
Keyword(s):  
Dna Polymerase ◽  
Dna Amplification ◽  
Geobacillus Stearothermophilus ◽  
Large Fragment ◽  
Dna Polymerase I ◽  
Polymerase I

scholarly journals Enhancement of Polymerase Activity of the Large Fragment in DNA Polymerase I from Geobacillus stearothermophilus by Site-Directed Mutagenesis at the Active Site

2016 ◽  
Vol 2016 ◽  
pp. 1-8 ◽  
Author(s):  
Yi Ma ◽  
Beilei Zhang ◽  
Meng Wang ◽  
Yanghui Ou ◽  
Jufang Wang ◽  
...  
Keyword(s):  
Dna Polymerase ◽  
Rapid Detection ◽  
Polymerase Activity ◽  
Site Directed Mutagenesis ◽  
High Price ◽  
Geobacillus Stearothermophilus ◽  
Large Fragment ◽  
Dna Polymerase I ◽  
Primary Diagnostics ◽  
Polymerase I

The large fragment of DNA polymerase I from Geobacillus stearothermophilus GIM1.543 (Bst DNA polymerase) with 5′-3′ DNA polymerase activity while in absence of 5′-3′ exonuclease activity possesses high thermal stability and polymerase activity. Bst DNA polymerase was employed in isothermal multiple self-matching initiated amplification (IMSA) which amplified the interest sequence with high selectivity and was widely applied in the rapid detection of human epidemic diseases. However, the detailed information of commercial Bst DNA polymerase is unpublished and well protected by patents, which makes the high price of commercial kits. In this study, wild-type Bst DNA polymerase (WT) and substitution mutations for improving the efficiency of DNA polymerization were expressed and purified in E. coli. Site-directed substitutions of four conserved residues (Gly310, Arg412, Lys416, and Asp540) in the activity site of Bst DNA polymerase influenced efficiency of polymerizing dNTPs. The substitution of residue Gly310 by alanine or leucine and residue Asp540 by glutamic acid increased the efficiency of polymerase activity. All mutants with higher polymerizing efficiency were employed to complete the rapid detection of EV71-associated hand, foot, and mouth disease (HFMD) by IMSA approach with relatively shorter period which is suitable for the primary diagnostics setting in rural and underdeveloped areas.

Site-specific modification of E. coli DNA polymerase I large fragment with pyridoxal 5'-phosphate

Biochemistry ◽  
1984 ◽  
Vol 23 (9) ◽  
pp. 2073-2078 ◽  
Author(s):  
Anup K. Hazra ◽  
Sevilla Detera-Wadleigh ◽  
Samuel H. Wilson
Keyword(s):  
Dna Polymerase ◽  
Large Fragment ◽  
Dna Polymerase I ◽  
Site Specific ◽  
E Coli ◽  
Specific Modification ◽  
Polymerase I

Sequencing with the large fragment of DNA polymerase I fromBacillus stearothermophilus

DNA Sequence ◽  
1991 ◽  
Vol 1 (3) ◽  
pp. 173-180 ◽  
Author(s):  
J. McClary ◽  
S. Y. Ye ◽  
G. F. Hong ◽  
F. Witney
Keyword(s):  
Dna Polymerase ◽  
Large Fragment ◽  
Dna Polymerase I ◽  
Polymerase I
Sign in / Sign up

Export Citation Format

Share Document