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Quantitative molecular hybridization with unfractionated, solubilized cells using RNA probes and polyacrylamide gel electrophoresis

1987; Elsevier BV; Volume: 167; Issue: 2 Linguagem: Inglês

10.1016/0003-2697(87)90180-1

1096-0309

Gary S. Firestein, Susan M. Gardner, W Roeder,

DNA and Nucleic Acid Chemistry

Molecular hybridization with RNA probes was performed on unfractionated cells solubilized in guanidine thiocyanate solutions. Unhybridized probe was digested with ribonuclease, and protected probe fragments were resolved by polyacrylamide gel electrophoresis (PAGE). Since the same medium was used both for solubilization of the cells and as the hybridization buffer, RNA purification was not required and the analysis of large numbers of samples was facilitated. Using this method, specificity is superior to dot blot analysis because the size of hybridized fragments is determined and the signal of the probe hybridized to target RNA is separated from the background by PAGE.