Selvamani, Ram Shankar Velur; Telaar, Maurice; Friehs, Karl; Flaschel, Erwin: Antibiotic-free segregational plasmid stabilization in Escherichia coli owing to the knockout of triosephosphate isomerase (tpiA). In: Microbial Cell Factories. Jg.13 H. 1. 2014
Inhalt
- Results
- Screening for an appropriate knockout gene
- Cloning of tpiA
- Application of the tpiA knockout in a recombinant production system
- Batch fermentation with strain JW3890-2-pFC1
- Batch fermentation with strain JW3890-2-pFC4
- Test of plasmid stabilization with tpiA in continuous culture
- Construction of a tpiA vector with altered downstream overlap
- Growth of the tpiA knockout strain on different media in the absence or presence of the complementing plasmid pJET-tpiA-Tart
- Analysis of plasmid stability
- Accumulation of methylglyoxal may lead to a toxic effect
- Antibiotic free cloning
- Discussion
- Conclusions
- Methods
- Cultivation
- Measurement of growth
- Determination of the specific growth rate
- Plasmid stability
- Determination of methylglyoxal concentration
- Determination of glycerol concentration
- Assay for β-Glucanase activity
- Cloning
- Sequencing
- Competing interests
- Authors’ contributions
- Acknowledgements
- References