Effect of Plasmid Copy Number and lac Operator Sequence on Antibiotic-Free Plasmid Selection by Operator-Repressor Titration in Escherichia coli

Autor:	Rocky M. Cranenburgh, Kathryn S. Lewis, Julian A. J. Hanak
Rok vydání:	2004
Předmět:	Plasmid preparation Genetics Physiology Cell Biology Biology Lac repressor Origin of replication Applied Microbiology and Biotechnology Biochemistry Microbiology Molecular biology PBR322 Plasmid maintenance Plasmid Low copy number Selectable marker Biotechnology
Zdroj:	Microbial Physiology. 7:197-203
ISSN:	2673-1673 2673-1665
DOI:	10.1159/000079828
Popis:	The Escherichia coli strain DH1lacdapD enables plasmid selection and maintenance that is free from antibiotics and selectable marker genes. This is achieved by using only the lac operator sequence as a selectable element. This strain is currently used to generate high copy number plasmids with no antibiotic resistance genes for use as DNA vaccines and for expression of recombinant proteins. Until now these have been limited to pUC-based plasmids containing a high copy number pMB1-derived origin of replication, and the principle lacO1 and auxiliary lacO3 operators. In this study we have shown that this system can also be used to select and maintain pBR322-based plasmids with the lower copy number pMB1 origin of replication, and that lacO1 alone or a palindromic version of lacO1 can provide a sufficient level of repressor titration for plasmid selection. This is advantageous for recombinant protein production, where low copy number plasmids are often used and plasmid maintenance is important. The degree of repressor titration due to these plasmids was measured using the natural lactose operon in E. coli DH1 as a model.
Databáze:	OpenAIRE
Externí odkaz:	https://explore.openaire.eu/search/publication?articleId=doi_________::6b510137c0939e981dce25f4dfbe67ad https://doi.org/10.1159/000079828 Zobrazit plný text záznamu