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Vol. 7, No. 4, 2004
Issue release date: September 2004
Section title: Research Article
J Mol Microbiol Biotechnol 2004;7:197–203
(DOI:10.1159/000079828)

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

Cranenburgh R.M. · Lewis K.S. · Hanak J.A.J.
Cobra Biomanufacturing Plc, The Science Park, Keele, Staffs., UK

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Article / Publication Details

First-Page Preview
Abstract of Research Article

Received: 3/15/2004
Accepted: 5/14/2004
Published online: 9/17/2004
Issue release date: September 2004

Number of Print Pages: 7
Number of Figures: 4
Number of Tables: 1

ISSN: 1464-1801 (Print)
eISSN: 1660-2412 (Online)

For additional information: http://www.karger.com/MMB

Abstract

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.

© 2004 S. Karger AG, Basel


  

Author Contacts

Rocky M. Cranenburgh
Cobra Biomanufacturing Plc
The Science Park
Keele ST5 5SP (UK)
Tel. +44 1782 714181, Fax +44 1782 799817, E-Mail rocky.cranenburgh@cobrabio.com

  

Article Information

Number of Print Pages : 7
Number of Figures : 4, Number of Tables : 1, Number of References : 29

  

Publication Details

Journal of Molecular Microbiology and Biotechnology

Vol. 7, No. 4, Year 2004 (Cover Date: Released September 2004)

Journal Editor: Saier, M.H., Jr. (La Jolla, Calif.)
ISSN: 1464–1801 (print), 1660–2412 (Online)

For additional information: http://www.karger.com/mmb


Article / Publication Details

First-Page Preview
Abstract of Research Article

Received: 3/15/2004
Accepted: 5/14/2004
Published online: 9/17/2004
Issue release date: September 2004

Number of Print Pages: 7
Number of Figures: 4
Number of Tables: 1

ISSN: 1464-1801 (Print)
eISSN: 1660-2412 (Online)

For additional information: http://www.karger.com/MMB


Copyright / Drug Dosage

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Drug Dosage: The authors and the publisher have exerted every effort to ensure that drug selection and dosage set forth in this text are in accord with current recommendations and practice at the time of publication. However, in view of ongoing research, changes in goverment regulations, and the constant flow of information relating to drug therapy and drug reactions, the reader is urged to check the package insert for each drug for any changes in indications and dosage and for added warnings and precautions. This is particularly important when the recommended agent is a new and/or infrequently employed drug.
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