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How to induce p53? - (Dec/16/2005 )

Hi Everybody!

I want to evaluate p53 activation in my cells by a western blotting. I would like to include a positive control, i.e. a lysates from cells where p53 was induced. Does anyone know how a protocol for p53 induction?

cheers

j

-josephine-

QUOTE (josephine @ Dec 16 2005, 04:50 PM)
Hi Everybody!

I want to evaluate p53 activation in my cells by a western blotting. I would like to include a positive control, i.e. a lysates from cells where p53 was induced. Does anyone know how a protocol for p53 induction?

cheers

j

Induction of p53 activity can be tested specifically by irradiation of your cells with X-ray (5Gy) and look for induction of p21 mRNA or protein or look at G1 arrest by FACS. The response to other types of radiation like UV is not solely dependent on p53.

-Theo22-

j
[/quote]
Induction of p53 activity can be tested specifically by irradiation of your cells with X-ray (5Gy) and look for induction of p21 mRNA or protein or look at G1 arrest by FACS. The response to other types of radiation like UV is not solely dependent on p53.
[/quote]


thanks, but im afraid i dont have the equipment for x rays in the lab. any other idea?

-josephine-

Hi

I would get a p53 inducible plasmid, tranfect, then induce and make lysates from the induced cells. You should be able to detect p53 directly by western.

Cheers,
Bob

-bob1-

QUOTE (josephine @ Dec 16 2005, 07:50 AM)
Hi Everybody!

I want to evaluate p53 activation in my cells by a western blotting. I would like to include a positive control, i.e. a lysates from cells where p53 was induced. Does anyone know how a protocol for p53 induction?

cheers

j



Treat your culture with doxorubicin overnight. It will induce p53.

-Soni-

etoposide 1-10 ┬ÁM overnight?

Seb_

-tryptofan-