We have developed a set of procedures which simultaneously solubilize, denature, and stabilize the RNA present in whole cells or in cytoplasmic extracts for direct analysis by gel electrophoresis and hybridization of blotted gels. Multiple samples of tissue culture cells and be prepared for electrophoresis in less than an hour. The number of cells that can be processed for whole cell RNA is limited only by the sensitivity of detection of specific RNAs using hybridization probes. Cell preparations with high levels of RNase, such as human lymphocytes and HL60, can be processed reliably. The method involves the stabilization of RNA in solubilized cells, cytoplasm or nuclei using various combinations of vanadyl ribonucleoside complex (VRC), bentonite, SDS, proteinase K, formaldehyde and heat, followed by resolution and specific detection of the RNA species on formaldehyde gels containing SDS. The transcripts of a number of different genes have been specifically detected using methodology to directly prepare and resolve the RNA from the cells of a variety of different tissue culture cell lines. We have recently extended the capability of this methodology to include the direct preparation and resolution of specific RNA's from the cells of solid tissues and tumors. At present, we are testing the ability of this technique to screen for and to assay the modulation of the level of messenger RNA transcribed for relevant genes, such as oncogenes.
