muramidase and cesium-chloride

A simple RNA isolation method was developed to purify bacterial RNAs from a large number of samples simultaneously in an hour. The method is based on boiling the cells in the presence of Triton X-100 and lysozyme, and then preferential RNA precipitation with ammonium acetate. There is no CsCl centrifugation required. For the nitrogen-fixing bacterium Klebsiella pneumoniae, the depression condition can be maintained during the cell-harvesting process. The intact isolated RNAs appeared to be free of protein, with a yield of 100 micrograms RNA from a 4-ml cell culture of 100 Klett units (10(9) cells/ml). Any DNA present was in a form that did not react with a nifH probe following Northern blotting to nitrocellulose (i.e., was not single-stranded).

Topics: Acetates; Blotting, Northern; Centrifugation, Density Gradient; Cesium; Chemical Precipitation; Chlorides; Detergents; Electrophoresis, Agar Gel; Klebsiella pneumoniae; Methods; Muramidase; Octoxynol; Polyethylene Glycols; RNA, Bacterial