TCF/LEF Luciferase Reporter Lentivirus (Wnt/β-catenin Signaling Pathway) are replication incompetent, HIV-based, VSV-G pseudotyped lentiviral particles that are ready to transduce almost all types of mammalian cells, including primary and non-dividing cells. The particles contain a firefly luciferase reporter under the control of TCF (T cell factor)/LEF (lymphoid enhancer factor)-responsive element located upstream of the minimal TATA promoter. The lentiviruses also transduce a puromycin selection gene (Figure 1). After transduction, activation of the Wnt/β-catenin signaling pathway in target cells can be monitored by assessing luciferase expression.
Figure 1. Schematic of the lenti-vector used to generate the TCF/LEF Luciferase Reporter Lentivirus (Wnt/β-catenin Signaling Pathway).
Two vials (500 µl x 2) of TCF/LEF Luciferase Reporter Lentivirus (Wnt/β-catenin Signaling Pathway) at ≥107 TU/ml. The titer will vary with each lot; the exact value is provided with each shipment.
The lentiviruses were produced from HEK293T cells. Supplied in medium containing 90% DMEM + 10% FBS. Virus particles can be packaged in custom formulations and produced at higher titers by special request, for an additional fee.
Background
The Wnt signaling pathway is a well described regulator of organism development and cell fate specification. Cellular pluripotency and differentiation of multiple lineages, including cardiomyocytes and neurons are modulated by this pathway. There are 19 Wnt ligands and 10 Frizzled receptors, which stimulate canonical as well as non-canonical Wnt pathways. In the canonical pathway, cell activation by a Wnt ligand leads to the stabilization and nuclear translocation of β-catenin and subsequent activation of TCF (T cell factor)/LEF (lymphoid enhancer factor) transcription factor. Dysfunction of these pathways can result in cancer and also type 2 diabetes. A deeper understanding of the role of this pathway and the development of new therapeutic strategies around it will open new clinical avenues.
References
Tian S., et al., Blood. 1994; 84(6):1760-1764. Zhong, Z., et al., Science. 1994; 264(5155):95-98.
Storage/Stability
Lentiviruses are shipped with dry ice. For long-term storage, it is recommended to store the lentiviruses at -80°C for up to 12 months from date of receipt. Avoid repeated freeze/thaw cycles. Titers can drop significantly with each freeze/thaw cycle.
Applications
Screen for activators or inhibitors of Wnt signaling pathway in transduced target cells.
Generate TCF/LEF reporter cell pools or stable cell lines following puromycin selection.
Shipping Temperature
-80°C
Notes
To generate a TCF/LEF luciferase reporter-expressing stable cell line, remove the growth medium 48 hours after transduction and replace it with fresh growth medium containing the appropriate amount of puromycin (as pre-determined from a killing curve, https://bpsbioscience.com/cell-line-faq), for antibiotic selection of transduced cells, followed by clonal selection.
The following Lentivirus Reporter Controls are available from BPS Bioscience to meet your experimental needs:
Negative Control Luciferase Lentivirus (#79578): Ready-to-transduce lentiviral particles expressing firefly luciferase under the control of a minimal promoter. The negative control is important to establish the specificity of any treatments and to determine the background reporter activity.
Renilla Luciferase Lentivirus (#79565): Ready-to-transduce lentiviral particles expressing Renilla luciferase under the control of a CMV promoter. The RLuc lentivirus can serve as an internal control to overcome sample-to-sample variability when performing dual-luciferase reporter assays.
Firefly Luciferase Lentivirus (#79692-G, #79692-H, #79692-P): Ready-to-transduce lentiviral particles expressing firefly luciferase under the control of a CMV promoter. It can serve as a positive control for transduction optimization studies.
Biosafety The lentiviruses are produced with the SIN (self-inactivation) lentivector which ensures self-inactivation of the lentiviral construct after transduction and integration into the genomic DNA of the target cells. None of the HIV genes (gag, pol, rev) will be expressed in the transduced cells, as they are expressed from packaging plasmids lacking the packing signal. Although the pseudotyped lentiviruses are replication-incompetent, they require the use of a Biosafety Level 2 facility. BPS recommends following all local federal, state, and institutional regulations and using all appropriate safety precautions.
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