3 replies to this topic

[clarem]

Hi,
Just wondering if anyone has any experience with transfecting large plasmids into HEK293 cells?
I am starting work with a plasmid of approximately 10kb and have been warned that I may have difficulties getting this into the cells.
If anyone has any protocols or advice, such as any particular transfection reagents that are good for transfection of large plasmids, I would be really grateful.
Thanks

[bob1]

I'm routinely doing 9 kb into 293.  You shouldn't have too much problem using either lipofectamine or fugene.  If you really struggle you may want to look at electroporation systems such as the Neon from Invitrogen/Life technologies.

[rmh999]

clarem, on 11 January 2011 - 03:25 AM, said:

Hi,
Just wondering if anyone has any experience with transfecting large plasmids into HEK293 cells?
I am starting work with a plasmid of approximately 10kb and have been warned that I may have difficulties getting this into the cells.
If anyone has any protocols or advice, such as any particular transfection reagents that are good for transfection of large plasmids, I would be really grateful.
Thanks

I've regularly transfected 13+ kb plasmids with Lipofectamine 2000 or Geneporter II into 293 cells and never had any real issues.

[apeters]

Hello,

Why not try nanoparticles - easy to do, can transfect large plasmids, and into difficult to transfect cell lines, to boot!  http://www.nanotherics.com

APeters