vurquidi at worf.molbiol.ox.ac.uk
Tue Apr 1 10:21:31 EST 1997
In Article <whc23-0104971105490001 at powercenter-604.physiol.cam.ac.uk>,
whc23 at cus.cam.ac.uk (Dr.W.H. Colledge) wrote:
>Does anyone have a simple, foolproof (preferably in a single Eppendorf
>tube) method for obtaining a cytoplasmic and nuclear extract from tissue
I use the following protocol:
Washed pellet of cells.
Resuspend in TNE- 1% NP-40 (or 0.5% NP-40)
(TNE = 10mM Tris-Cl (pH 8.00)/100 mM NaCl/1 mM EDTA)
Leave on ice for 5 min with occasional agitation.
Nuclei are removed from the lysate by centrifugation
(1500 xg for 15 min 4 C)
Works always well. The nuclei pellet is compact and very white.
If it is not, probably the lysis was incomplete.
More information about the Methods