Transfection didnt work after repeating it. - (Aug/31/2010 )
After leaving this for a while, i came back to it. I used cells at a lower passage, and tried different transfection methods, and optimatisations.
As it stands, i have my control GFP transfecting in fine into the cells.
And the cells look reasonably healthy afterwards.
But i'm not getting anythign with my target gfp.
I do see some small lumps of green fluorescence outside of the cells however.
What are these?
Is it a case that the plasmid is too big to go in, or what could be wrong here?
What is the your target GFP? Is it a protein fusion betwen GFP and your protein of interst? Is so, that protein willbe subject to localization and/or degradation depending on the fusion partner. The other possibility is that there is an issue in the genetic construct.
Hope this is helpful.