help: plant microsome preps

Q. Shang qshang at u.washington.edu
Thu Jul 25 14:28:50 EST 1996


Hi there,

I have been trying to make microsome preps out of cultured plant cells,
without any luck so far (no microsome pellets after centrifugation at
100kg).  The biggest problem I had was to break the cells; I tried warring
blender, mortar and pestle, sonication,french press, all failed to break
the cell walls (judging from observations under microscope).  I was
finally able to lyse the cells by treating with cellulase for hours and
then grinding in the mortar, sometimes with glass beads.  I suspect that
this treatment may be too rough, i.e.,may have damaged the ER membrane. I
thought about grinding cells in liquid nitrogen, but wonder if that would
be deleterious to the membrane proteins I'm trying to isolate.  I'm quite
at a loss here, so any suggestions would be most appreciated.

Tanya

-----------------------------
Tanya Shang
Department of Biochemistry
Box 357350
University of Washington
Seattle, WA 98195
qshang at u.washington.edu
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