Rapid Purification of 70 S RNA from Media of Cells Producing RNA Tumor Viruses

Abstract

Media from cells producing RNA tumor viruses, when treated with sodium dodecyl sulfate and polyvinyl sulfate, yield 70 S RNA as the major species binding oligo(dT)-cellulose. The procedure described for purifying 70 S RNA requires no special equipment and is suitable for rapidly processing large quantities of media or for purifying RNA from commercially available virus, with a 5- to 10-fold higher yield than was obtained using existing methods.

Endogenous RD-114 virus genome expression in malignant tissues of domestic cats

“…Virus-infected cell lines were grown in phosphate-free Eagle minimum essential medium and 10% dialyzed fetal bovine serum for 12 h. The cells were then labeled for four 12-h periods with the above media supplemented with 100 ,LCi of carrierfree [3uPiphosphoric acid per ml (27). The 32P-labeled viral RNA was isolated from the virions released in the last 12-h culture fluids using oligodeoxythymidylate-cellulose as described by Smith et al (36). The poly(A)-containing RNA solution was treated with SDS-Pronase in the presence of 0.1 mg ofE.…”

Primate type-C virus nucleic acid sequences (woolly monkey and baboon types) in tissues from a patient with acute myelogenous leukemia and in viruses isolated from cultured cells of the same patient

“…Unlabeled 70S viral RNA was purified according to Wu et al (26), except that the buffer used for glycerol gradient centrifugation was 0.1 M LiCl, 0.01 M Tris-HCI at pH 7.4, 0.001 M EDTA, and 0.2% sodium dodecyl sulfate. Cell DNA was purified (27); 3H-labeled 70S viral RNA was obtained (15,28); [3H]DNA copies of viral RNA (cDNA) were prepared under conditions specified earlier (3) and then were purified according to a modification of the procedure of Baxt and Spiegelman (5). The cDNA-containing solution was extracted with phenol-chloroform at pH 9 (27), and nucleic acids were made 0.2 M in NaCl and precipitated with 2 volumes of EtOH.…”