IMPROVED METHOD FOR PURIFICATION OF VIRAL-RNA FROM FECAL SPECIMENS FOR ROTAVIRUS DETECTION

An improved procedure to recover and purify double-stranded RNA (dsRNA ) from fecal specimens is described. Guanidine isothiocyanate, hydroxy apatite, and cetyltrimethylammonium bromide were used to extract and p urify rotavirus dsRNA from fecal specimens. The method is very efficie nt and easy to perform, and precludes the use of toxic substances such as phenol, chloroform, and Freon. It yields RNA free of enzymatic inh ibitors, permitting its detection by reverse transcription-polymerase chain reaction assays. In addition, it was demonstrated that during in itial clarification of the fecal suspension, the pellet must be washed at least twice to avoid massive losses of virus, viral protein, or vi ral nucleic acid retained in the solid debris.