Difference between revisions of "BACTH"
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see a scheme of the principle: [http://www.euromedex.com/shop/popup_image.php?pID=208357&osCsid=a63d2e53d8afa86ab50e58493b07b400 picture] | see a scheme of the principle: [http://www.euromedex.com/shop/popup_image.php?pID=208357&osCsid=a63d2e53d8afa86ab50e58493b07b400 picture] | ||
| − | The system consists of four | + | The system consists of four [[plasmid]]s and one E. coli strain (BTH101) |
# pKT25: C-terminal fusion of the target protein to the T25 fragment of adenylate cyclase | # pKT25: C-terminal fusion of the target protein to the T25 fragment of adenylate cyclase | ||
| Line 14: | Line 14: | ||
# pUT18C: C-terminal fusion of the target protein to the T18 fragment of adenylate cyclase | # pUT18C: C-terminal fusion of the target protein to the T18 fragment of adenylate cyclase | ||
| − | E. coli BTH101 is a cya mutant that requires plasmid-derived adenylate cyclase activity for the activation of the ''lac'' operon. | + | E. coli BTH101 is a cya mutant that requires [[plasmid]]-derived adenylate cyclase activity for the activation of the ''lac'' operon. |
References: | References: | ||
Revision as of 12:01, 16 April 2009
Bacterial Adenylate Cyclase-based two hybrid system
Allows the detection of in vivo protein-protein interactions by restoring the activity of the Bordetella pertussis adenylate cyclase in E. coli reporter strains.
Active adenylate cyclase results in the expression of the lacZ gene which is detected by blue colonies on plates containing X-Gal.
see a scheme of the principle: picture
The system consists of four plasmids and one E. coli strain (BTH101)
- pKT25: C-terminal fusion of the target protein to the T25 fragment of adenylate cyclase
- pKT25-N: N-terminal fusion of the target protein to the T25 fragment of adenylate cyclase
- pUT18: N-terminal fusion of the target protein to the T18 fragment of adenylate cyclase
- pUT18C: C-terminal fusion of the target protein to the T18 fragment of adenylate cyclase
E. coli BTH101 is a cya mutant that requires plasmid-derived adenylate cyclase activity for the activation of the lac operon.
References:
- Karimova, G., Fayolle, C., Gmira, S., Ullmann, A., Leclerc, C., and Ladant, D. (1998) Charge-dependent translocation of Bordetella pertussis adenylate cyclase toxin into eukaryotic cells: implication for the in vivo delivery of CD8(+) T cell epitopes into antigen-presenting cells. Proc Natl Acad Sci USA 95, 12532-12537. PubMed
- Claessen, D., Emmins, R., Hamoen, L. W., Daniel, R. A., Errington, J., and Edwards, D. H. (2008) Control of the cell elongation-division cycle by shuttling of PBP1 protein in Bacillus subtilis. Mol Microbiol 68, 1029-1046. PubMed
The BACTH is commercially available from EUROMEDEX
