Characterization of cold-active uracil-DNA glycosylase from Bacillus sp HJ171 and its use for contamination control in PCR SCIE SCOPUS

Cited 3 time in WEB OF SCIENCE Cited 4 time in Scopus
Title
Characterization of cold-active uracil-DNA glycosylase from Bacillus sp HJ171 and its use for contamination control in PCR
Author(s)
Kim, Gun A.; Lee, Mi Sun; Sun, Younguk; Lee, Byung Doo; Il Lee, Jong; Lee, Jung-Hyun; Kwon, Suk-Tae
KIOST Author(s)
Lee, Jung Hyun(이정현)
Publication Year
2008-10
Abstract
In this study, the gene encoding Bacillus sp. HJ171 uracil-DNA glycosylase (Bsp HJ171 UDG) was cloned and sequenced. The Bsp HJ171 UDG gene consists of a 738-bp DNA sequence, which encodes for a protein that is 245-amino-acid residues in length. The deduced amino acid sequence of the Bsp HJ171 UDG had a high sequence similarity with other bacterial UDGs. The molecular mass of the protein derived from this amino acid sequence was 27.218 kDa. The Bsp HJ171 UDG gene was expressed under the control of a T7lac promoter in the pTYB1 plasmid in Escherichia coli BL21 (DE3). The expressed enzyme was purified in one step using the Intein Mediated Purification with an Affinity Chitin-binding Tag purification system. The optimal temperature range, pH, NaCl concentration, and KCl concentration of the purified enzyme was 20-25 degrees C, 8.0, 25 and 25 mM, respectively. The half-life of the enzyme at 40 degrees C and 50 degrees C were approximately 131 and 45 s, respectively. These heat-labile characteristics enabled Bsp HJ171 UDG to control carry-over contamination in the polymerase chain reaction product (PCR) without losing the PCR product.
ISSN
0175-7598
URI
https://sciwatch.kiost.ac.kr/handle/2020.kiost/4443
DOI
10.1007/s00253-008-1585-0
Bibliographic Citation
APPLIED MICROBIOLOGY AND BIOTECHNOLOGY, v.80, no.5, pp.785 - 794, 2008
Publisher
SPRINGER
Subject
COD GADUS-MORHUA; ESCHERICHIA-COLI; N-GLYCOSIDASE; EXPRESSION; CLONING; PURIFICATION; POLYMERASE; SEQUENCE
Keywords
Bacillus sp; HJ171 (Bsp HJ171); uracil-DNA glycosylase (UDG); Bsp HJ171 UDG; psychrophile; carry-over contamination
Type
Article
Language
English
Document Type
Article
Publisher
SPRINGER
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