Transfection of cells by viral DNA - (Oct/10/2005 )
We want to transfect viral DNA into cells. The viral DNA is a year old. What would be a better option- cloning the DNA or reamplifying it by PCR before using it for transfection.
Also what DNA concentration is required for efficient transfection?
Since the DNA is year old are there any factors that are needed to be taken into account?
Your response will be highly appreciated.
What virus is it? Is it in a plasmid? If it was stored @ minus 20 it should be fine.
i agree with TAP 14. The important point is not the conc of DNA but the relative quantity of DNA in regard of number of cells. The quality of DNA should be ok.
For amplification, PCR is quicker.
Its a polyoma virus and it was stored in TE/water at -20. would 1ug be ok...or should it be amplified before transfection??
It should be perfect. DNA is very stable at -20 especially in TE. The concentration you use is going to vary depending on the transfection protocol and reagent you decide to use. Some of the lipofectamine reagent for example can use very small amounts of DNA and have a very high transfection efficiency in certain cell lines. You should figure out what protocol people have used for your cells and go with that.