I have transfected a cell with a bicistronic vector "Gene of interest and GFP" for transient expression in HEK293T. The cells are transfected successfully, as they continue to survive in Zeocin selection pressure, moreover there are some GFP positive population, however after few week of maintenance under Zeocin selection there is no GFP cells ,i.e cells without GFP expression is surviving and distributed throughout the flask .

I assume CMV promoter cold be silenced by epigenetic mechanism .. Can some one confirm my assumption. If there is any other reason , kindly let me know..

My gene cassette is like this

CMV promoter -T7 Pomoter- Transgene of interest -chimeric intron -IRES-eGFP- pAsignal-SV40 & EM7 promoter -Bleo R gene.

Note: This question might resemble with already discussed question. Only difference is my protein of interest (GFP) expressing cells are seen in the beginning of Zeocin maintenance,

https://www.researchgate.net/post/Why_are_my_cells_not_expressing_my_protein_of_interest_after_transfection

https://www.researchgate.net/post/Transfection_without_expression

Thanks ..

Regards

Suresh