Elsevier

Analytical Biochemistry

Volume 484, 1 September 2015, Pages 180-182
Analytical Biochemistry

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Single cell-level detection and quantitation of leaky protein expression from any strongly regulated bacterial system

https://doi.org/10.1016/j.ab.2015.06.011Get rights and content

Abstract

Extremely low levels of “leaky” expression of genes in bacterial protein expression systems can severely curtail cell viability when expressed proteins are toxic. A general method for sensitive detection of such expression is lacking. Here, we present a method based on microscopic visualization of a fluorescent “reporter” protein (RFP–HU-A) constructed by fusing red fluorescent protein (RFP) to the N-terminus of a nucleoid-associated, histone-like DNA-binding protein, HU-A. Localization of RFP–HU-A within nucleoids facilitates detection, quantitation, and characterization of leaky expression at the single-cell level.

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Acknowledgments

K.A. thanks the Universities Grants Commission for a doctoral research fellowship. P.G. thanks IISER Mohali and the Ministry of Human Resource Development, Govt. of India, for a Center of Excellence grant (MHRD-14-0064) in Protein Science, Design and Engineering.

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