Edited by mexemi, 07 October 2004 - 07:28 AM.
Help with stable cell lines
Posted 07 October 2004 - 07:22 AM
Posted 11 October 2004 - 09:25 PM
as to your amount of selection antibiotic, it doesn't really matter what you use as long as you have healthy cells and enough of a selection pressure at all times. I personally like hitting cells with a high dose, and when only a few are left I decrease the antibiotic.
You say that [QUOTE]Many cells died but there are still so many cells so I cannot pick isolated colonies with a Gilson,
Are you sure that you have stably transfected cells, are you using enough G418? Normally only a few cells survive, so that you should be able to pick colonies.
Posted 13 October 2004 - 01:53 AM
Rember to verify that your clone(s) express what you have transfected the cells with, and hopefully that they express it more than the pool of cells you isolated the clone from. This can be done with RT-PCR og Northern blot.
I think the selection method is determined on the basis of what vector you use.
Posted 14 October 2004 - 05:16 AM