English
 
User Manual Privacy Policy Disclaimer Contact us
  Advanced SearchBrowse

Item

ITEM ACTIONSEXPORT
  Effects of a recombinant gene expression on ColE1-like plasmid segregation in Escherichia coli

Popov, M., Petrov, S. A., Nacheva, G., Ivanov, I., & Reichl, U. (2011). Effects of a recombinant gene expression on ColE1-like plasmid segregation in Escherichia coli. BMC Biotechnology, 11(1), 18. doi:10.1186/1472-6750-11-18.

Item is

Basic

show hide
Item Permalink: http://hdl.handle.net/11858/00-001M-0000-0013-8D49-C Version Permalink: http://hdl.handle.net/11858/00-001M-0000-002C-8C33-5
Genre: Journal Article

Files

show Files
hide Files
:
564645_bmc_bio_2011.pdf (Publisher version), 2MB
Name:
564645_bmc_bio_2011.pdf
Description:
-
Visibility:
Public
MIME-Type / Checksum:
application/pdf / [MD5]
Technical Metadata:
Copyright Date:
-
Copyright Info:
This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/2.0), which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

Locators

show

Creators

show
hide
 Creators:
Popov, M.1, Author              
Petrov, S. A.1, Author
Nacheva, G.1, Author
Ivanov, I.1, Author              
Reichl, U.2, 3, Author              
Affiliations:
1Bulgarian Academy of Sciences Institute of Molecular Biology "Roumen Tsanev" Sofia, Bulgaria, ou_persistent22              
2Otto-von-Guericke-Universität Magdeburg, ou_1738156              
3Bioprocess Engineering, Max Planck Institute for Dynamics of Complex Technical Systems, Max Planck Society, ou_1738140              

Content

show
hide
Free keywords: -
 Abstract: BACKGROUND:Segregation of expression plasmids leads to loss of recombinant DNA from transformed bacterial cells due to the irregular distribution of plasmids between the daughter cells during cell division. Under non-selective conditions this segregational instability results in a heterogeneous population of cells, where the non-productive plasmid-free cells overgrow the plasmid-bearing cells thus decreasing the yield of recombinant protein. Amongst the factors affecting segregational plasmid instability are: the plasmid design, plasmid copy-number, host cell genotype, fermentation conditions etc. This study aims to investigate the influence of transcription and translation on the segregation of recombinant plasmids designed for constitutive gene expression in Escherichia coli LE392 at glucose-limited continuous cultivation. To this end a series of pBR322-based plasmids carrying a synthetic human interferon-gamma (hIFNgamma) gene placed under the control of different regulatory elements (promoter and ribosome-binding sites) were used as a model.RESULTS:Bacterial growth and product formation kinetics of transformed E. coli LE392 cells cultivated continuously were described by a structured kinetic model proposed by Lee et al. (1985). The obtained results demonstrated that both transcription and translation efficiency strongly affected plasmid segregation. The segregation of plasmid having a deleted promoter did not exceed 5% after 190 h of cultivation. The observed high plasmid stability was not related with an increase in the plasmid copy-number. A reverse correlation between the yield of recombinant protein (as modulated by using different ribosome binding sites) and segregational plasmid stability (determined by the above model) was also observed.CONCLUSIONS:Switching-off transcription of the hIFNgamma gene has a stabilising effect on ColE1-like plasmids against segregation, which is not associated with an increase in the plasmid copy-number. The increased constitutive gene expression has a negative effect on segregational plasmid stability. A kinetic model proposed by Lee et al. (1985) was appropriate for description of E. coli cell growth and recombinant product formation in chemostat cultivations. © 2011 Popov et al; licensee BioMed Central Ltd. [accessed 2013 November 14th]

Details

show
hide
Language(s): eng - English
 Dates: 2011
 Publication Status: Published in print
 Pages: -
 Publishing info: -
 Table of Contents: -
 Rev. Method: Peer
 Identifiers: eDoc: 564645
DOI: 10.1186/1472-6750-11-18
 Degree: -

Event

show

Legal Case

show

Project information

show

Source 1

show
hide
Title: BMC Biotechnology
Source Genre: Journal
 Creator(s):
Affiliations:
Publ. Info: BioMed Central
Pages: - Volume / Issue: 11 (1) Sequence Number: - Start / End Page: 18 Identifier: ISSN: 1472-6750
CoNE: /journals/resource/111000136906066