This is an open-ended methods question for anyone out there. We're at the point in our lab where we're about to implant various cell populations into inbred hosts, with the hope of verifying that these cells are surviving and incorporating into normal tissue. Survival times range from 4 weeks to 26 weeks.
What methods does your lab use to do this? We've considered lentiviral transduction and transfection with GFP/luciferase expressing plasmids.
We plan to create stable cell lines expressing GFP/mCherry under cell-specific promoters by infecting these cells with lentiviral particles (in collaboration with Dr Howell's lab). We hope to get these constructs made by the end of the summer. Our ubiquitous promoter (EF1alpha) driving mCherry worked well but it messed up our cell lines -- unexpected regulation of marker genes using RTPCR. So, we will have to screen through the lines to look for cells that behave like the uninfected.
ReplyDeleteThat sounds like a tough task. I'm curious to know how this goes once you get to that point.
ReplyDeleteDid you make the construct yourself or did you order it from a company?