High yield nuclear extract preparation

Thomas thomas.korosecNOthSPAM at univie.ac.at.invalid
Mon Dec 20 05:23:43 EST 1999


I am currently preparing nuclear extracts from mouse liver
used in DNA footprinting reactions. I am isolating nuclei
according to a modifyed method described by Widnal et.
al.(Homogenization of the tissue in 300 mM sucrose following
centrifugation through a 2.2 M sucrose cushion). This gives
miscroscopically pure nuclei in a acceptable yield. The
nuclear extract is then prepared by ammonium sulphate
precipitation followed by dialysis (published by Parker et.
al.). My problem is a very very low yield (about 10
micrograms of nuclear extract from 10 grams liver) at the
end, whereas published papers decribe yields being 1000
times higher (10 mg extract from 10 g tissue). Has anybody
recommendations how to improve yields.


Thanks !


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