Dear colleagues,
Please share your experience in using the psiCHECK-2 vector (Promega) for luciferase assay.
I have experience with pGL3 and pRL vectors before and I always got results that the Firefly (test) raw luciferase data is higher than the Renilla raw data (endogenous control).
In the psiCHECK-2 vector, Renilla is expressed under SV40_promotor/enchancer and is a test luciferase, whereas Firefly is expressed under HSV-TK promoter and is an endogenous control. Therefore, I expected that in psiCHECK-2 Renilla's raw expression would be higher than Firefly’s. However, in my first experiments with empty psiCHECK-2, I found that raw Renilla signal is about 10 times lower than Firefly, so I have low value of Rluc/Fluc ratio (about 0.1).
Is it suitable for psiCHECK-2 vector and what is the reason for such observations? What are the approximate raw values you have when using psiCHECK-2?
To measure luciferase activity I use Dual-Glo® Luciferase Assay System (Promega).
Thank you in advance for your feedback.
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