TetR-controlled recombination-based in vivo expression technology (TRIVET) vector

Contributors

Inventor Institute
Stefan Schild University of Graz
SKU: 160861
Product description: This system is based upon recombination-based in vivo expression technology (RIVET) and allows the detection of transient gene silencing events even in a subset of a complex, heterogenous bacterial population. It requires all three components; the tpc reporter cassette, pTRIVET and pTRIVET1. It can be used as a random approach to identify in vivo repressed genes during intestinal colonization and as a specific approach to study conditional transcriptional silencing of a gene of interest. The protocol for which can be found at PMID: 33659431.
Alternate name: TRIVET
Additional notes: Vibrio cholerae gene expression
Cat. #: 160861
Tool sub type: Reporter
Research Fields: Microbiology
Target: Cholera
Bacterial resistance: pTRIVET – ampicillin, tpc reporter cassette – chloramphenicol
Selectable markers: PhoA activity
Application notes: This system is based upon recombination-based in vivo expression technology (RIVET) and allows the detection of transient gene silencing events even in a subset of a complex, heterogenous bacterial population. It requires all three components; the tpc reporter cassette, pTRIVET and pTRIVET1. It can be used as a random approach to identify in vivo repressed genes during intestinal colonization and as a specific approach to study conditional transcriptional silencing of a gene of interest. The protocol for which can be found at PMID: 33659431.
References:

33659431

Cakar et al. 2018. Proc Natl Acad Sci U S A. 115(10):E2376-E2385. PMID: 29463743.

Images

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A resolvase-based screen to identify in vivo repressed genes. (A) Schematic illustration of the genetic components of the TetR-controlled in vivo expression technology (TRIVET). Chromosomal sequences are highlighted in light gray; tetR-phoA-cat (tpc) cassette in black; integrated pTRIVET in dark gray; and the res cassette parts in open shapes. The tpc cassette is integrated into a V. cholerae hypothetical geneX via Tn10 mutagenesis; resulting in a transcriptional fusion of tetR and phoA to the chromosomal promoter of geneX. Subsequently; pTRIVET is integrated downstream of lacZ. Thus; the expression level of TetR via the geneX promoter dictates the expression of TnpR; which catalyzes the excision of the res cassette. The IS10 elements and constitutive cat promoter of the tpc cassette; the TetR-controlled tnpR; mobilization (mob); origin of replication (oriR6K); and ApR (bla) regions of pTRIVET; as well as the gene for KmR (neo); SucS (sacB); and the target sites of resolvase (res) of the res cassette are indicated. (B) Shown are the effects of iron availability on resolution frequency (red) and alkaline phosphatase (PhoA) activity (black) of strain Vc_res1_TRIVET irgA

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