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Biochem Biophys Res Commun. 2018 Feb 12;496(3):820-825. doi: 10.1016/j.bbrc.2018.01.147. Epub 2018 Jan 31.

Application of the SSB biosensor to study in vitro transcription.

Author information

1
School of Biosciences, University of Kent, Canterbury, CT2 7NJ, UK.
2
School of Biosciences, University of Kent, Canterbury, CT2 7NJ, UK. Electronic address: c.toseland@kent.ac.uk.

Abstract

Gene expression, catalysed by RNA polymerases (RNAP), is one of the most fundamental processes in living cells. The majority of methods to quantify mRNA are based upon purification of the nucleic acid which leads to experimental inaccuracies and loss of product, or use of high cost dyes and sensitive spectrophotometers. Here, we describe the use of a fluorescent biosensor based upon the single stranded binding (SSB) protein. In this study, the SSB biosensor showed similar binding properties to mRNA, to that of its native substrate, single-stranded DNA (ssDNA). We found the biosensor to be reproducible with no associated loss of product through purification, or the requirement for expensive dyes. Therefore, we propose that the SSB biosensor is a useful tool for comparative measurement of mRNA yield following in vitro transcription.

KEYWORDS:

Biosensor; Gene expression; Myosin; RNA polymerase; SSB; Transcription; mRNA

PMID:
29378185
PMCID:
PMC5811048
DOI:
10.1016/j.bbrc.2018.01.147
[Indexed for MEDLINE]
Free PMC Article

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