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Description: This is a luciferase reporter of beta-catenin-mediated transcriptional activation. The idea for the construct is from Hans Clevers lab, who designed the original TOPflash. However, this version has a much higher signal/noise ratio. In HEK cells, maximal activation of this reporter is ~100-fold (activation by Wnt) up to ~1,000-fold (activation by phosphorylation mutants of beta-catenin). The appropriate control plasmid is clone M51, Super8XFOPflash, which has mutant TCF/LEF binding sites.
This construct was made by Ajamete Kaykas in the Moon lab. The backbone is the pTA-Luc vector of Clontech, which provides a minimal TA viral promoter driving expression of the firefly luciferase gene (see company publications for details). 16 TCF/LEF binding sites were cloned into the Mlu1 site of this vector (16 copies of: AGATCAAAGGgggta, with TCF/LEF binding site in CAP letters, and a spacer in lower case, separating each copy of the TCF/LEF site).
Notes for Use: Select clones with Ampicillin.
Reference: Made by Ajamete Kaykas, citation Science 308: 826-833 (2005)