Make your personal pGreenFire 2.zero reporter vector or use as a detrimental management
With the pGreenFire 2.zero mCMV Cloning & Unfavorable Management Lentivector & Virus (pGF2-mCMV-rFluc-T2A-GFP-mPGK-Puro), you may make the most of our sturdy pGreenFire 2.zero lentivector know-how to create your personal transcriptional response component (TRE) reporter or use pGreenFire 2.zero mCMV as-is as a detrimental management.
With the pGreenFire 2.zero mCMV Cloning & Unfavorable Management Lentivector, XhoI and NheI websites are positioned upstream of a minimal CMV promoter (mCMV) so you may clone in your personal TREs. Upon activation, the TREs and mCMV promoter collectively drive co-expression of crimson firefly luciferase and GFP so you may quantitatively measure transcriptional exercise utilizing each fluorescence and luciferase exercise.
Alternatively, you need to use the vector as-is as a detrimental management for any undertaking utilizing pGreenFire 2.zero lentivectors (see information under).
What makes our next-gen pGreenFire 2.zero vectors even higher than different TRE reporter vectors is the good design, which provides in a constitutive choice cassette for steady cell line era whereas minimizing interference with the upstream TRE. By utilizing a weak/reasonable mPGK promoter to drive the antibiotic choice marker (puromycin resistance) and thoroughly arranging the conditional reporter genes, the choice marker is reliably expressed with out compromising conditional expression of rFLuc and GFP.
NEW PRODUCT ALERT!
SBI is worked up to announce the launch of our subsequent era of pGreenFire signalling pathway reporters! We’ve upgraded these standard lentivectors with a intelligent design that permits dependable era of steady cell traces and have additionally changed the traditional luciferase reporter with crimson firefly luciferase, which opens up the potential of performing a dual-spectral luciferase assay and delivers larger sensitivity for in vivo functions than typical luciferase.
SBI Lentiviral Applied sciences
pGreenFire Fundamentals
For those who’re not acquainted with our pGreenFire reporters, each 1.zero and a couple of.zero share an analogous core performance—transcriptional response parts (TREs) are positioned upstream of a minimal CMV promoter (mCMV) and the pGreenFire luciferase-T2A-GFP co-expression cassette. Within the absence of transcriptional activation, the mCMV promoter has negligible exercise leading to little- to no- luciferase exercise or GFP fluorescence (Determine 1). Nonetheless, upon activation of the TREs, corresponding to in response to the addition of an inducer, the TREs plus the mCMV promoter drive expression of each luciferase and GFP in a dose-dependent trend (Determine 1). The result’s the flexibility to quantitatively measure pathway activation utilizing luciferase exercise or whereas imaging utilizing GFP.
As with all of our pGreenFire 2.zero lentivectors, the GreenFire cassette now consists of crimson firefly luciferase (rFLuc), a T2A co-expression component, and GFP. The swap to rFLuc opens up the potential of performing a dual-spectral luciferase assay and likewise delivers larger sensitivity for in vivo functions than typical luciferase.
