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isolating bacillus spores - (Feb/02/2006 )

I have grown several different bacillus cultures and am in process of isolating the spores from the vegetative cells in the final culture. My final wash of the spores involves putting them thru a 5 micron filter. However, some of the cultures are still showing a large amount of vegetative cells, and I need a cleaner batch of spores. I have read about heating the samples at 80 degrees C for ten minutes to lyse the vegetative cells. Does anyone else have any other suggestions or is this the best way to obtain clean spore samples?

Any suggestions appreciated,


i have purified proteins from the spore surfacen they way i lysed them was to grow them in spitzens of schaeffers medium to the relevant stage;
collect cells and resuspend in 50mM tric-HCl with 1mg/ml lysozyme, 2mM DTT, 1 mM PMSF, 1 mM EDTA. then incubated the suspension at 4oC for 4 hours with shaking.

active protein was recovered in spore fraction

alternatively you can pop the cells by passing them through a french press if you have one its quicker than cold lysis!!!