Measurement of Transcription, Translation, and Other Enzymatic Processes During Cell-Free Expression Using PERSIA

We developed the PERSIA technique with an interest in quantifying proteins as they are being produced during a cell-free synthesis reaction. A short 6-amino acid sequence added to a protein of interest reacts with a fluorogenic reagent (ReAsH), yielding a measure of protein concentration in close to...

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Bibliographic Details
Published inMethods in molecular biology (Clifton, N.J.) Vol. 2433; p. 169
Main Authors Wick, Scott, Carr, Peter A
Format Journal Article
LanguageEnglish
Published United States 2022
Subjects
Persia
Protein Biosynthesis
RNA, Messenger - genetics
Persia
PERSIA
Protein quantitation
Fluorescent label
Translation
Transcription
Cell-free synthesis
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Summary:We developed the PERSIA technique with an interest in quantifying proteins as they are being produced during a cell-free synthesis reaction. A short 6-amino acid sequence added to a protein of interest reacts with a fluorogenic reagent (ReAsH), yielding a measure of protein concentration in close to real time. We combine this measurement with simultaneous fluorescent detection of mRNA production, quantifying both transcription and translation. Alternatively, we combine simultaneous measurement of protein synthesis and that protein's enzymatic activity. We have found these simple capabilities enabling for multiple applications, including sequence-structure-function studies and target-specific assessment of drug candidate compounds.
ISSN:1940-6029
DOI:10.1007/978-1-0716-1998-8_10