Application

Set of transfection grade T cell factor (TCF) reporter plasmids for use in TOPFlash and FOPFlash wnt/b-catenin activity assays.

Components

FOPflash (mutant TCF binding sites) (Cat.# 21-169)TOPflash (TCF Reporter Plasmid) (Cat.# 21-170)

Disclaimer

Unless otherwise stated in our catalog or other company documentation accompanying the product(s), our products are intended for research use only and are not to be used for any other purpose, which includes but is not limited to, unauthorized commercial uses, in vitro diagnostic uses, ex vivo or in vivo therapeutic uses or any type of consumption or application to humans or animals.

General description

FOPflash: Transfection grade T cell factor (TCF) reporter plasmid containing two full and one incomplete copy of the TCF binding site (mutated) followed by three copies in the reverse orientation, upstream of the Thymidine Kinase (TK) minimal promoter and Luciferase open reading frame. This plasmid serves as a negative control to TOPflash which contains wild type TCF binding sites.

TOPflash: Transfection grade T-cell factor (TCF) reporter plasmid containing two sets (with the second set in the reverse orientation) of three copies of the TCF binding site (wild type) upstream of the Thymidine Kinase (TK) minimal promoter and Luciferase open reading frame. FOPflash containing mutated TCF binding sites is also available as a negative control.

The Wnt family encodes secreted glycoproteins implicated in cell growth regulation, cell fate determination, organogenesis and oncogenesis. Inferred from genetic analysis of wingless (wg) signaling in Drosophila, in mammalian cells, Wnts signal through the Frizzled family of receptors, to Dishevelled. Dishevelled down regulates glycogen synthase kinase-3 (GSK-3) activity. This results in the stabilization and accumulation of cytosolic -catenin, a GSK-3 substrate, which when phosphorylated by GSK-3, is targeted for ubiquitin-mediated proteolysis. -catenin then translocates to the nucleus, where in complexes with members of the Tcf/LEF family of transcription regulators, activates transcription of Tcf-responsive genes. This Tcf-reporter plasmid, along with FOPFLASH (Cat. # 21-169) enables quantitation of Wnt/Wg signaling in cells transfected with these constructs.

Legal Information

UPSTATE is a registered trademark of Merck KGaA, Darmstadt, Germany

Physical form

FOPflash, Catalog # 21-169, Lot # 30860, 5.5 kb. 5 µg affinity purified DNA eluted and aseptically packaged in 50 µL of 10 mM Tris-HCl, pH 8.0, 1 mM EDTA. Frozen solution.

TOPflash, Catalog # 21-170, Lot # 27752, 5.5 kb. 5 µg affinity purified DNA eluted and aseptically packaged in 50 µL of 10 mM Tris-HCl, pH 8.0, 1 mM EDTA. Frozen solution.

Quality

Routinely evaluated by DNA Sequence Analysis to confirm the identity of TCF binding sites.

Storage and Stability

Stable for 2 years at -20°C from date of shipment. For maximum recovery of product, centrifuge the original vials after thawing and prior to removing the cap.