world

[PING CHEN]

Dear bionetters,
	I have been trying to prepare nuclear extracts from promonocytic cell
lines (U1, U937) for a while and have not been very successful. I used 
Dignam's protocol or the modified version of the protocols that other people 
used successfully for making nuclear extracts. My problem is that I could not
get a majority of the cells to lyse without breaking the nuclei as judged by 

(1) whole cells were still present after homogenization with a glass Dounce 
    homogenizer - after over 20 strokes;

(2) the "nuclear pellets" get very viscous when I tried to resuspend them, 
    which I take as an indication that DNA was out in solution. 

The protocols seem straight forward and many people reference them in the
litereature.  I do not know if I am missing some important hints.  I would
appreciate it if anybody who reads this message and has experience and is 
willing to help a desperate grad.student.

Ping Chen

pchen at welchlink.welch.jhu.edu