fungal protein extraction problem

Maura Meade meade at unity.ncsu.edu
Fri Mar 6 00:58:25 EST 1998


Hi folks

	I've been having a problem with extracting protein from Cochliobolus
heterostrophus.  I lyophilize mycelium, grind it in liquid N2 and then
in extraction buffer (0.1M phosphate buffered saline, 10% glycerol, 4mM
B-mercaptoethanol, 1% triton-X-100  and assorted proteinase
inhibitors).  I spin this at 6-10K Gs and collect the supernatant for
protein determination and SDS-PAGE.  When I do this with another fungus
(Cercospora kikuchii) it works fine, meaning I see bands ranging from
high to low MW on a SDS-PAGE gel.  With Cochliobolus extracts however I
get a few high MW bands, a few low MW bands, and nothing in between.
This doesn't seem to be a degradation problem. The only difference I
notice between the extracts from these two fungi is that the
Cochliobolus extracts are viscous (polysaccharides?).  Does anyone have
any suggestions how to remedy this?  Thanks

Mark




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