How to wash pellet before RNA isolation - (May/01/2007 )
I am optimizing the RNA isolation for Rhizobium etli. When I centrifuge a culture in stationary phase, the pellet is not so clear. There is a cloud of polysaccharides above it. How can I get rid of it??
I stabilize the RNA by adding 1/5 volume 95/5 ethanol/phenol to the culture and dropping the falcon in liquid nitrogen. After thawing I centrifuge at 4750 rpm (maximum speed) for 30 min at 4 degrees. But the pellet is not clear.
I've tried washing it with TE-buffer (pH 8) to the pellet. But I'm not certain that the polysaccharides dissolve in the buffer. Will the RNA isolation remain intact? Anyone an idea?
if the cells are intact, wash them in PBS.
If not, after a first separation by acid phenol chloroform IAA, do an other PCI followed by chloroform only treatment.
Should allow the most part of polysacchatrides to get over.