cotransfect or bicistronic

Alistair Forrest zzalforr at fox.uq.net.au
Wed Mar 15 18:04:59 EST 2000


We've played around with cotransfections and the IRESpuro bicistronic
vector.

With cotransfections you are right in the assumption that a large
percentage of your GFP expressing cells will also express your gene of
interest, however not all of them will, you can tip things in your favour
by using relatively little EGFP plasmid comapred to your GOI vector. Those
that fluoresce are more likely to contain the GOI compared to a
transfection with equimolar amounts (maybe try 4:1 or 9:1). This will give
you fewer green cells but they are more likely to contain your GOI.

Alternatively use the IRES vector and avoid all the fuss. The IRES puro
vector can allow you to select for your overexpressors within a day or two
(much quicker than G418 or hygro).

good luck,
	Al




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