Construction of a reporter system for bifidobacteria using chloramphenicol acetyltransferase and its application for evaluation of promoters and terminators.

ABSTRACT: A reporter assay system is an essential tool for investigating gene expression mechanisms. In the case of bifidobacteria, several convenient and sensitive reporter systems have been developed. Here, we developed a new reporter system for bifidobacteria using the chloramphenicol acetyltransferase gene (cat) from Staphylococcus aureus. This enzyme stoichiometrically produced free CoA-SH, which was analyzed quantitatively with Ellman's test using 2-nitrobenzoic acid (DTNB). The 2-nitro-5-thiobenzoate (TNB^2-) produced showed a strong yellowish color with maximum absorbance at 412 nm. We also constructed a new pBCMAT plasmid series for CAT assays in bifidobacteria to evaluate promoters and terminators. Analyses using promoters from Bifidobacterium longum NCC2705

SUBMITTER: Kozakai T 

PROVIDER: S-EPMC8099631 | biostudies-literature | 2021

REPOSITORIES: biostudies-literature