Clonetech Retro-X Tet-on system - (Jan/12/2015 )

Hi-

I'm confused! I have 2 vectors-

pRetroX-Tet-On Advanced   &

pRetroX-Tight-Pur (with my gene of interest)

My question: Tell me if I'm doing this right---I was planning on taking regular 293T cells and doing an infection into 293T (like I do for lenti)- with VSVg, delta, pRetroX-Tet-On Advanced vector using Fugene--harvest the virus and infect my cells, MCF-7 and generate a G418-resistant stable line that expresses the transactivator.

I was then going to do another infection into 293T like before but use pRetroX-Tight-Pur (with my gene of interest). And then infect the MCF-7/pRetroX-Tet-On Advanced line with sup that I generated---Does that sound right?

It seems odd that I'm using the same infection protocol for retro that I have used for lenti??? 

Are infection protocols similar. I remember from years ago--people would use Phoenix cells than 293T

The first process is correct. 

First is select stable clone which stable express rtTA, and next step is transfect CMVtet-Target gene to rtTA stable clone.

Would be a waste of time. 

You also can construct double promoter vector which express rtTA and target gene at same time.