Posttranscriptional regulation of gene expression by small noncoding RNAs (sRNAs) is an important control mechanism that modulates bacterial metabolism, motility, and pathogenesis. Using the bacterial carbon storage regulator/regulator of secondary metabolism (Csr/Rsm) system, we here describe an E. coli-based cell-free translation assay that allows a quantitative analysis of translation regulation by ncRNAs and their corresponding translation repressor proteins. The assay quantifies the translation of chloramphenicol acetyltransferase in cell-free expression reactions that contain defined amounts of ncRNA and repressor protein. We demonstrate our protocol with a comparative translation activation analysis of the RsmX, RsmY, and RsmZ sRNAs from Pseudomonas protegens, which reveals a superior efficacy of RsmZ over RsmX and RsmY.
CITATION STYLE
Michel, E., Duss, O., & Allain, F. H. T. (2018). An integrated cell-free assay to study translation regulation by small bacterial noncoding RNAs. In Methods in Molecular Biology (Vol. 1737, pp. 177–195). Humana Press Inc. https://doi.org/10.1007/978-1-4939-7634-8_11
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