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pre-prepararation of competent cells - (Mar/24/2010 )

yaster day i cultured e.coli cells in 5 ml media then incubate over night at 37C for activation after freezing in liquid nitrogen,
to day i sub cultured the cells in 50 ml media, the problem which i filled in is that i incubate the cells over night at 37C instead of 18 C!!
i totally forgot to adjust the incubator to 18 C.
i want to know , if i can complete my steps to prepare competent cells, or i to repeat the steps from the bigenning ? .
thanks a lot

-aidaelshaer-

I've never heard of overnight at 18C.
What method are you using? I'm intrigued!

-leelee-

I think he means that he incubated them at 18C and not at 37C... ?

If not then it is indeed strange.

But if you just incubated at 18 instead of 37, well yeah, your cells will not have grown "perfectly", but you can still try to use them.

The circumstances aren't perfect, but try to use them anyway.

-pito-

pito on Mar 25 2010, 05:31 AM said:

I think he means that he incubated them at 18C and not at 37C... ?

If not then it is indeed strange.

But if you just incubated at 18 instead of 37, well yeah, your cells will not have grown "perfectly", but you can still try to use them.

The circumstances aren't perfect, but try to use them anyway.



in general method we make cells to grow under fasting condition to make its wall weak, by incubation at 18 C for 16 to 20 hrs,
or growing at 37C for 3 hrs,
any way i solved this problem by taking 10 ml of the culture and ubculture it at 37C or room temperature for 2-3 hrs.

thanks alot.

-aidaelshaer-