How many plasmid DNA copies per cell? - (Feb/26/2007 )

Hi everyone,

I am trying to transfect primary human skeletal muscle cells, which isn´t that easy. But, I am wondering, how many copies of plasmid DNA do I need (per cell) to see an increase in expression of my cDNA insert (which is already in the genome but I want to increase its expression)? Did anybody count their successful transfection in terms of copie numbers?
I am curious to see your answers.

Thanks in advance!

humpback-whale

the increase in expression of the cDNA insert will depend not only on the number of copies of the plasmid in the cell but on the strength of the promotor used to drive the expression in the first place.

quantifying transgene expression by copy number of plasmid wont tell you very much, just because the plasmid is present doesnt mean its expressing anything (promotor silencing etc.) to quantify transgene expression is easy using rtPCR