ChIP in mouse tissues, How to crosslink ? - (Mar/08/2007 )

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Hi,
I wonder if anyone has done ChIP on tissues? I would like to do ChIP in livers from mouse. Anyone who knows how to crosslink and how to extract the protein-DNA complexes ?

Thanks in advance!

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Others have posted on this topic so you should be able to find a few protocols. Here's one from my lab:

Take tissue sample from -80C and crush it till you get fragments about 1cm cubed (err on the small side) using a mortar and pestle cooled with liquid nitrogen (however you go about it you want to end up with frozen fragments about 1cm in dimension).

Quickly add one fragment to 1ml of 1% formaldehyde in PBS at room temp and quickly mince with scizzors to very small pieces. The tissue should incubate in the formaldehyde for 20min from the time you start mincing.

Spin the fragments down at high speed in a refrigerated microfuge for 1min. Aspirate supe and add 125mM glycine in PBS. Incubate for about 5min at room temp. Spin again, aspirate, and then wash twice with ice cold PBS (keep on ice from this point on).

From this point, treat as you normally would for ChIP samples. We use 1ml of lysis buffer for this much tissue but I would suggest splitting the lysate into two 500ul aliquots before sonicating.

Good luck.

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