Microbial nar-GFP cell sensors reveal oxygen limitations in highly agitated and aerated laboratory-scale fermentors.

Microbial Cell Factories
Jose R GarciaWilliam E Bentley

Abstract

Small-scale microbial fermentations are often assumed to be homogeneous, and oxygen limitation due to inadequate micromixing is often overlooked as a potential problem. To assess the relative degree of micromixing, and hence propensity for oxygen limitation, a new cellular oxygen sensor has been developed. The oxygen responsive E. coli nitrate reductase (nar) promoter was used to construct an oxygen reporter plasmid (pNar-GFPuv) which allows cell-based reporting of oxygen limitation. Because there are greater than 109 cells in a fermentor, one can outfit a vessel with more than 109 sensors. Our concept was tested in high density, lab-scale (5 L), fed-batch, E. coli fermentations operated with varied mixing efficiency - one verses four impellers. In both cases, bioreactors were maintained identically at greater than 80% dissolved oxygen (DO) during batch phase and at approximately 20% DO during fed-batch phase. Trends for glucose consumption, biomass and DO showed nearly identical behavior. However, fermentations with only one impeller showed significantly higher GFPuv expression than those with four, indicating a higher degree of fluid segregation sufficient for cellular oxygen deprivation. As the characteristic time for GFPuv ...Continue Reading

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Citations

Oct 11, 2011·Biotechnology Letters·Isaac Vizcaino-CastonTim W Overton
Feb 27, 2009·Microbial Cell Factories·Frank DelvignePhilippe Thonart
Aug 26, 2014·Current Opinion in Biotechnology·Karen M Polizzi, Cleo Kontoravdi
Oct 23, 2010·Biotechnology and Bioengineering·Maëlle DouairePascal Loubière
Aug 18, 2009·Biotechnology and Bioengineering·Alvaro R LaraWouter A van Winden
Mar 19, 2014·Proceedings of the National Academy of Sciences of the United States of America·Jonathan W KotulaPamela A Silver
Nov 30, 2018·Molecular Systems Biology·Felix MoserChristopher A Voigt
Feb 26, 2020·Microbial Cell Factories·Lei ZhaoXiaoyuan Wang
Mar 20, 2018·Journal of Industrial Microbiology & Biotechnology·Mitchell S Weisenberger, Tara L Deans
Mar 16, 2018·Bioprocess and Biosystems Engineering·Anna-Lena Heins, Dirk Weuster-Botz
May 8, 2021·Microbial Biotechnology·Xinyi WanBaojun Wang

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