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Investigation of plasmid-induced growth defect in Pseudomonas putida

Paper ID Volume ID Publish Year Pages File Format Full-Text
22661 43369 2016 7 PDF Available
Title
Investigation of plasmid-induced growth defect in Pseudomonas putida
Abstract

•Maintenance of pMiS1 plasmid affects P. putida cell growth.•Newly identified low copy rep mutants of the pBBR1 replicon lead to wildtype-like growth.•Kanamycin resistance gene and transcription factor encoding rhaR gene are causal for growth defect.

Genetic engineering in bacteria mainly relies on the use of plasmids. But despite their pervasive use for physiological studies as well as for the design and optimization of industrially used production strains, only limited information about plasmid induced growth defects is available for different replicons and organisms. Here, we present the identification and characterization of such a phenomenon for Pseudomonas putida transformants carrying the pBBR1-derived plasmid pMiS1. We identified the kanamycin resistance gene and the transcription factor encoding rhaR gene to be causal for the growth defect in P. putida. In contrast, this effect was not observed in Escherichia coli. The plasmid-induced growth defect was eliminated after introduction of a mutation in the plasmid-encoded rep gene, thus enabling construction of the non-toxic variant pMiS4. GFP reporters construct analyses and qPCR experiments revealed a distinctly lowered plasmid copy number for pMiS4, which is probably the reason for alleviation of the growth defect by this mutation. Our work expands the knowledge about plasmid-induced growth defects and provides a useful low-copy pBBR1 replicon variant.

Keywords
Plasmid induced growth defect; Metabolic burden; Pseudomonas putida; Plasmid copy number; pBBR1 replicon; Rep; KanR
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Investigation of plasmid-induced growth defect in Pseudomonas putida
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Publisher
Database: Elsevier - ScienceDirect
Journal: Journal of Biotechnology - Volume 231, 10 August 2016, Pages 167–173
Authors
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Subjects
Physical Sciences and Engineering Chemical Engineering Bioengineering
Get Full-Text Now
Don't Miss Today's Special Offer
Price was $35.95
You save - $31
Price after discount Only $4.95
100% Money Back Guarantee
Full-text PDF Download
Online Support
Any Questions? feel free to contact us