Make your private pGreenFire 2.zero reporter vector or use as a detrimental administration
With the pGreenFire 2.zero mCMV Cloning & Unfavorable Administration Lentivector & Virus (pGF2-mCMV-rFluc-T2A-GFP-mPGK-Puro), chances are you’ll benefit from our sturdy pGreenFire 2.zero lentivector know-how to create your private transcriptional response part (TRE) reporter or use pGreenFire 2.zero mCMV as-is as a detrimental administration.
With the pGreenFire 2.zero mCMV Cloning & Unfavorable Administration Lentivector, XhoI and NheI web sites are positioned upstream of a minimal CMV promoter (mCMV) so chances are you’ll clone in your private TREs. Upon activation, the TREs and mCMV promoter collectively drive co-expression of crimson firefly luciferase and GFP so chances are you’ll quantitatively measure transcriptional train using every fluorescence and luciferase train.
Alternatively, you want to use the vector as-is as a detrimental administration for any endeavor using pGreenFire 2.zero lentivectors (see data underneath).
What makes our next-gen pGreenFire 2.zero vectors even larger than totally different TRE reporter vectors is the nice design, which offers in a constitutive alternative cassette for regular cell line period whereas minimizing interference with the upstream TRE. By using a weak/cheap mPGK promoter to drive the antibiotic alternative marker (puromycin resistance) and completely arranging the conditional reporter genes, the selection marker is reliably expressed with out compromising conditional expression of rFLuc and GFP.
SBI Lentiviral Applied sciences
GreenFire Fundamentals
For many who’re not acquainted with our pGreenFire reporters, every 1.zero and a few.zero share an identical core efficiency—transcriptional response components (TREs) are positioned upstream of a minimal CMV promoter (mCMV) and the pGreenFire luciferase-T2A-GFP co-expression cassette. Throughout the absence of transcriptional activation, the mCMV promoter has negligible train resulting in little- to no- luciferase train or GFP fluorescence (Decide 1). Nonetheless, upon activation of the TREs, akin to in response to the addition of an inducer, the TREs plus the mCMV promoter drive expression of every luciferase and GFP in a dose-dependent pattern (Decide 1). The result is the pliability to quantitatively measure pathway activation using luciferase train or whereas imaging using GFP.
As with all of our pGreenFire 2.zero lentivectors, the GreenFire cassette now consists of crimson firefly luciferase (rFLuc), a T2A co-expression part, and GFP. The swap to rFLuc opens up the potential of performing a dual-spectral luciferase assay and likewise delivers bigger sensitivity for in vivo features than typical luciferase.
NEW PRODUCT ALERT!
SBI is labored as much as announce the launch of our subsequent period of pGreenFire signalling pathway reporters! We’ve upgraded these customary lentivectors with a clever design that allows reliable period of regular cell traces and have moreover modified the normal luciferase reporter with crimson firefly luciferase, which opens up the potential of performing a dual-spectral luciferase assay and delivers bigger sensitivity for in vivo features than typical luciferase.
