Rose-Hulman Undergraduate Research Publications
 

Document Type

Article

Publication Date

8-30-2017

First Advisor

Irene Reizman

Abstract

Microbial biosensors can be used to provide information about the cells’ environment in large-scale fermentations. In this project an oxygen sensitive biosensor is being developed in Escherichia coli to determine what kind of conditions cells are growing in: aerobic or anaerobic. This project specifically studies expression from the fumarate and nitrate reductase (FNR) promoter (PFNR) under aerobic and anaerobic conditions. In fluorescence experiments, the expression levels of green fluorescent protein (GFP) were used to quantify the effectiveness of the PFNR in the DH5α, MG1655 and BL21 strains of E. coli. The negative control of the experiments, which are the E. coli strains without a GFP plasmid, showed very high levels of background fluorescence until the emission and excitation wavelengths of the fluorescence plate reader were adjusted. Overall, the PFNR expressed more GFP under anaerobic conditions in the DH5α and BL21 strains. The results of the MG1655 strain indicated that the strain has the fnr- genotype and did not draw any definite conclusions of PFNR. Another oxygen sensitive promoter plasmid, pKVS-vgb-GFP, was constructed using circular polymerase extension cloning (CPEC) to be tested in the future. A positive control plasmid, pKVS-J23101-GFP, was also constructed using CPEC that should not produce as high of an amount of GFP relative to the current positive control plasmid. Once verified as the correct plasmid size, the new positive control can be used in future fluorescence experiments. Acknowledgements I wish

Comments

RHURP 2017-02

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