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trobule with double stable transfection - (Dec/02/2007 )

Hey everyone,
I've been trying to do a double stable transfection with G418 and hygromycin (used both g418 and hygromycin at the same time). I 've ran a kill curve on my 293 cells for both antibiotics and used the corrosponding concentration. But i'm having a hard time getting any cells that survive after 2 weeks. I'm not sure what the problem is. Anyone have any tips or experience with double stable transfections. I think there is something wrong with the hygromycin part of the trasnfection. I was previously able to do a single stable transfection using the G418. The problem occurs with I preform teh G418 and hygromycin selection together.

Thanks,
joe

-veryslowjoe-

QUOTE (veryslowjoe @ Dec 2 2007, 06:33 PM)
Hey everyone,
I've been trying to do a double stable transfection with G418 and hygromycin (used both g418 and hygromycin at the same time). I 've ran a kill curve on my 293 cells for both antibiotics and used the corrosponding concentration. But i'm having a hard time getting any cells that survive after 2 weeks. I'm not sure what the problem is. Anyone have any tips or experience with double stable transfections. I think there is something wrong with the hygromycin part of the trasnfection. I was previously able to do a single stable transfection using the G418. The problem occurs with I preform teh G418 and hygromycin selection together.

Thanks,
joe


if in parallel double stable transfection is too difficult, I would try to do it sequently, first select a G418 stable then a hygromycin stable transfectant

-The Bearer-