Hi everyone,
I am using Dharmacon Trans-Lentiviral shRNA packaging kit with Calcium Phosphate Transfection Reagent to pack my GIPZ-shRNA of interest into viral particles. The manual says that "gag-pro and tat-rev are under control of the conditional tetracycline-responsive promoter element (TRE), limiting expression of these viral components strictly to cells expressing the tetracycline transactivator." Also, pTLA1-TOFF expression vector in the packaging mix contains the tetracycline-controlled transactivator (Tet-Off), which is required for the expression of gag-pro and tat-rev. So, it looks like the system has everything it needs to produce the viral particles. Now, I am using 293 FT cells as the packaging cell line. Although I did not add any doxycycline to the growth medium (since the system is Tet-Off), I did not manage to produce any virus. I could not find any information about the characteristics of these cells. Does anyone have any information regarding this cell line and this system? Did anyone have any problems regarding the virus production with this system?
Thanks so much.
Hi Eda, in our lab we are also using the GIPZ and TRIPZ from dharmacon but we are not using their packaging system. We are using pPAX.2 and pMD2.G from addgene . producer cells 293T lenti-X. It works for us
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