Ribonucleic Acid from Reovirus as Seen in Protein Monolayers by Electron Microscopy

Abstract

The ribonucleic acid of reovirus was extracted with 2 M sodium perchlorate solution and spread by the protein monolayer technique. Areas of the monolayer were transferred to support films, rotary shadowed, and observed in the electron microscope. Filaments of RNA obtained by extraction prior to spreading were similar in appearance and in distribution of contour lengths (0.2 to 1.2 μ) to those obtained by phenol extraction of the virus. Most of the filaments resulting from extraction of the virus suspension during spreading on a sodium perchlorate solution, however, were longer than 1 μ. The lengths of the longest filaments exceeded the 5 μ length predicted from chemical data for one single piece of complementary-stranded RNA in the reovirus particle. The short filaments, 1.2 μ and less in length, fell into a tri-modal pattern of length distribution with peaks at 0.35 μ, 0.60 μ and 1.10 μ. These shorter lengths probably resulted from breakage of the intact RNA during the extraction procedures. The consistently observed pattern of length distribution suggests that they represent relatively stable subunits of the molecule. Sodium perchlorate extracted reovirus RNA was thermally denatured in formaldehyde prior to spreading by the protein monolayer technique. Length distributions and relative numbers of filaments in the peaks of the tri-modal distribution pattern were similar to those found for unheated material when extracted prior to spreading. This similarity indicates that heating subsequent to extraction produced no further filament breakage. The thin, kinky appearance of the heated filaments, and the appearance of congruent pairs, indicated that heating had separated the strands of the complementary-stranded RNA subunits.