Can Sonication break Yeast for Protein Purification?

WANGHO at UCBEH.SAN.UC.EDU WANGHO at UCBEH.SAN.UC.EDU
Sun Feb 26 16:16:02 EST 1995


Hi there!

I am trying to purify the protein from the yeast.  My protein is a nuclear
protein.  As I know, 75% of the protein is in the nuclei.  The Glass-Bead
Disruption does no work very well for me.  I usually expressed my protein in
E.coli., which can be easily broken by Sonication.  But most protocal is
foucused on the Glass Beads.  Why not Sonication?  I am going to try it.  Is it
a bad idea?

In addition, RNA is always a trouble in my purification.  I believe there is a
lot of RNA (maybe tRNA) in my extract.  This might due to that the MW of my    
protein is around 120 kD and I have to take longer to get me protein.  Because
the protein crude extract will be used for cross-linking before the SDS-PAGE, 
the RNA gives me trouble to do the cross-linking reaction.  I am thinking about
the addition of RNase.  Does anyone try RNase before?  Any suggestion about how
much and how long for the RNase use?

Thanks.

Hong



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