I use lentiviral vectors (invitrogen pLenti6.3/TO/V5-DEST) for transgenesis of mESCs, but it is extremely inefficient in comparison with somatic cells.
My mESC media is DMEM/F12+20%KOSR+LIF+R2i (SB431542 and PD0325901) which have been developed in the Royan Institute (publication DOI: 10.1007/s12015-011-9306-y and 10.1007/s12015-013-9473-0). Lentiviral particles are concentrated and work efficiently on other mouse and human cell types.
I have the same issue. Like you, I have highly efficient transduction of somatic cells. I narrowed the issue down to 1) CMV driven transgene expression, and 2) issues with the IRES element driving expression of the selection marker. Separated expression of transgene and selection marker using PGK or EF1A promoters solved the problem.
Best of luck.
Thank you for your advise. I think it can bee the case for my experiment too. In my vectors transgene expression (including GFP in my control vector) is CMV-driven and the selection marker (blasticidin resistance) is expressed with a SV40 promoter.
But just to make sure:
Is it possible to compare the rate of genomic integration in somatic and ES cells with realtimePCR? (I mean to measure the copy number of transgene per genome, using an endogenous gene as a normalizer) Then I can say that whether it is because of the promoter or integration inefficiency.
It should be possible to do the real-time experiment, but Southern blottting would be a better (and more commonly used) approach.
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