Record Details

An alternative approach for screening active <i>Bam </i><span style="mso-bidi-font-weight:bold">HI<b> </b>variants: Overexpression in <span style="font-size:12.0pt;font-family:"Times New Roman"; mso-fareast-font-family:"Times New Roman";mso-ansi-language:EN-IN;mso-fareast-language: EN-IN;mso-bidi-language:AR-SA" lang="EN-IN">T-7 RNA polymerase based system</span></span>

NOPR - NISCAIR Online Periodicals Repository

View Archive Info
 
 
Field Value
 
Title An alternative approach for screening active Bam HI variants: Overexpression in T-7 RNA polymerase based system
 
Creator Acharya, Asha S
Roy, Kunal B
 
Description 303-308
The type II restriction endonuclease. BamHI , has been overexpressed in
E. coli by cloning the BamHI gene in frame with an E.
coli Ribosome Binding Site (RBS) under the T7 promoter of an E. coli expression vector pRSET
A. The expression level of BamHI
endonuclease using this construct was found to be higher than that
reported of the overexpressing clone pAEK 14. Our overexpressing clone, pAABRw in
BL21 cell in presence of BamHI
methylase in pMAP6 following induction with IPTG yields about 9.2x106  units per gram wet cell paste. In vivo activity
of the recombinant endonuclease could be confirmed by the SOS induction assay
in JH 139 cells even in the absence of T7 polymerase and cognate BamHI methylase because of leaky expression
in E. coli. This provides an alternate way to screen the active
endonuclease and its variants.
 
Date 2012-12-25T18:43:31Z
2012-12-25T18:43:31Z
2001-10
 
Type Article
 
Identifier 0975-0959 (Online); 0301-1208 (Print)
http://hdl.handle.net/123456789/15318
 
Language en_US
 
Rights CC Attribution-Noncommercial-No Derivative Works 2.5 India
 
Publisher NISCAIR-CSIR, India
 
Source IJBB Vol.38(5) [October 2001]