Document Detail


Development of a simple cell lysis method for recombinant DNA using bacteriophage lambda lysis genes.
MedLine Citation:
PMID:  18176547     Owner:  NLM     Status:  MEDLINE    
Abstract/OtherAbstract:
In this study, we describe the development of a simple and efficient method for cell lysis via the insertion of a bacteriophage lambda lysis gene cluster into the pET22b expression vector in the following order; the T7 promoter, a gene for a target protein intended for production, Sam7 and R. This insertion of R and Sam7 into pET22b exerted no detrimental effects on cellular growth or the production of a target protein. The induction of the T7 promoter did not in itself result in the autolysis of cells in culture but the harvested cells were readily broken by freezing and thawing. We compared the efficiency of the cell lysis technique by freezing and thawing to that observed with sonication, and determined that both methods completely disintegrated the cells and released proteins into the solution. With our modification of pET22b, the lysis of cells became quite simple, efficient, and reliable. This strategy may prove useful for a broad variety of applications, particularly in experiments requiring extensive cell breakage, including library screening and culture condition exploration, in addition to protein purification.
Authors:
Boyun Jang; Yuna Jung; Dongbin Lim
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Publication Detail:
Type:  Journal Article; Research Support, Non-U.S. Gov't    
Journal Detail:
Title:  Journal of microbiology (Seoul, Korea)     Volume:  45     ISSN:  1225-8873     ISO Abbreviation:  J. Microbiol.     Publication Date:  2007 Dec 
Date Detail:
Created Date:  2008-01-07     Completed Date:  2008-06-17     Revised Date:  -    
Medline Journal Info:
Nlm Unique ID:  9703165     Medline TA:  J Microbiol     Country:  Korea (South)    
Other Details:
Languages:  eng     Pagination:  593-6     Citation Subset:  IM    
Affiliation:
Department of Bioinformatics and Life Science, Soongsil University, Seoul, Republic of Korea.
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MeSH Terms
Descriptor/Qualifier:
Bacteriolysis*
Bacteriophage lambda / genetics*,  metabolism
Cloning, Molecular / methods
DNA, Recombinant / genetics
Electrophoresis, Polyacrylamide Gel
Escherichia coli / cytology,  genetics,  metabolism
Freezing
Microbial Viability / genetics
Plasmids / genetics
Sonication
Viral Proteins / genetics*,  metabolism
Chemical
Reg. No./Substance:
0/DNA, Recombinant; 0/Viral Proteins

From MEDLINE®/PubMed®, a database of the U.S. National Library of Medicine


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