transfecting TOPO vector - can it be done? (Jun/15/2009 )

hi all.

I have my desired DNA cloned inside TOPO cloning vector. Is it possible to transfect this vector into mammalian cells? Has anybody performed this? Did it go ok?

Thanks!!!

-cupidstunt-

You can transfect any DNA in - but does it have a promoter for expression?

-nofx-

Yes it is possible, and yes it should go OK. If you haven't done transfections before, you need to look at transfection reagents such as fugene or lipofectamine.

Note that both linear and circular DNA will transfect, but empty un-ligated TOPO vectors may not transfect well because of the topo-isomerase at the cloning sites.

-bob1-

Hi,

It dosent have a promoter, how can it be expressed??

-cupidstunt-

If it doesn't have a promoter it might still express, but at a very low level - probably not worth looking for. What vector are you using?

-bob1-

If it doesn't have a promoter it might still express, but at a very low level - probably not worth looking for. What vector are you using?

-bob1-

bob1 on Jun 24 2009, 01:35 AM said:

If it doesn't have a promoter it might still express, but at a very low level - probably not worth looking for. What vector are you using?

well its this gene which we have bought, it has come in TOPO vector. We are trying to clone it out and put it in pCDNA vector. Its seems to be difficult one for some reason.

-cupidstunt-