I have an insert ( ORF 6,9kb long) that I want to transfect in a human colon cancer cell line.
I initially wanted to do with lentivirus, since I could have a stable expression of my gene and potentially insert larger inserts than I could do with a regular lipofectamine transfection. However, I've read the lentivirus has an insert limit, around 9kb, which is the size of its genome, that it would be possible to obtain efficient titers. My insert + the lenti vector (tet on inducible system) would sum around 13kb.
I have also considered Adenovirus, however I couldnt find adenovirus plasmis with an inducible expression that could also fit my insert ( also got a limit of aprox 5kb).
Any ideas that how I could possibly (preferably stable and inducible) express my gene in a human cell line?
Have you thought of microinjection? 13 KB should be doable with it I believe. The transfection efficiency will be high and the expression will be stable. The main disadvantages of microinjection will be the requirement of specialised equipment and a limited number of cells you can microinject in one go.
- Badges
- Science method