34. THE RIBONUCLEIC ACIDS OF VIRUSES 297 



of virus RNA was essentially the same as that for Mengo and West Nile encephalitis 

 viruses. 



Alexander et al. lb7 - lbS - 159 isolated RNA from purified type I and type II polio 

 virus which was grown in either monkey kidney or human amnion cells in monolayer. 

 Osmotic shock and dodecyl sulfate treatment of the virus did not yield detectable 

 amounts of infectious RNA, but treatment of the virus with phenol, in the cold, 

 yielded infectious RNA. The infectivity of the RNA solution was tested on HeLa 

 and human amnion cell monolayers on which the virus forms plaques. The RNA 

 preparations produced confluent cytopathogenic effects on these monolayers if an 

 undiluted preparation was tested directly after the phenol treatment. The cyto- 

 pathogenic effect occurs even when the period of contact is only 30 seconds. When 

 diluted with physiological saline solutions, the infectivity is greatly decreased, but 

 after dilution in hypertonic salt solutions (1 M NaCl with 0.04 M phosphate buffer, 

 pH 7.2) a high degree of infectivity is maintained. In appropriate environments, the 

 degree of infectivity (plaque-forming units) of the RNA is proportional to the dilu- 

 tion factor. The infectivity is almost 1% of the infectivity of the virus and remains 

 stable for 6-8 weeks when stored at — 70°C. 



The RNA preparations showed a UV spectrum characteristic for nucleic acid. The 

 protein content was not determined. Proteolytic enzymes (chymotrypsin and papain) 

 had no effect on the infectivity. Ribonuclease destroyed the activity and so did 

 normal monkey serum which contains ribonuclease. Deoxyribonuclease, normal 

 globulin, and polio-immune gamma globulin from rabbit serum, all of which showed 

 no ribonuclease activity, had no effect upon the infectivity. 



d. Ribonucleic Acid from Other Viruses 



Franklin et at. 168 prepared an infectious nucleic acid fraction from brains of mice 

 infected with mouse encephalomyelitis virus (ME virus) by cold and hot phenol 

 treatment. In contrast to the greatly reduced infectivity of EEE virus-RNA pre- 

 pared with hot phenol, the infectivity of ME virus-RNA obtained with hot phenol 

 treatment was the same as that of RNA isolated from ME virus-infected tissues by 

 cold phenol extraction. The procedure used for Mengo encephalitis virus was also 

 used here. This consists of ribonuclease treatment of homogenates of infected tissues 

 prior to phenol extraction. The results of this experiment indicate that the infectious 

 RNA is present in the cell in a form which is resistant to ribonuclease, probably as 

 intact virus. This is in contrast to the situation with EEE virus. Infectious RNA was 

 also isolated from virus which had been partially purified using the fluorocarbon 

 Arcton 63. This agent does not affect the infectivity of ME virus, whereas an exten- 

 sive treatment with Arcton 63 does destroy the infectious principle in EEE virus-in- 

 fected tissues which are the source of infectious EEE virus-RNA. 169 It was also 

 possible to isolate infectious RNA from a highly purified preparation of ME virus. 

 In all cases, the infectious ME virus-RNA was assayed by intracerebral injection in 

 mice. Discrete plaques could be produced on monolayers of Earle's L-cells but were 

 not suitable for quantitative tests. Huppert and Sanders 170 employed the phenol 

 method to prepare an infectious RNA component from cells of the Krebs 2 mouse 

 ascites carcinoma which had been infected with murine encephalomyocarditis virus. 

 The authors indicate that intact virus cannot be the sole source of infectious RNA 



16 » R. M. Franklin, E. Wecker, and C. Henry, Virolgy 7, 220 (1959). 



169 R. M. Franklin and E. Wecker, unpublished data (1959). 



170 J. Huppert and F. K. Sanders, A'a^re 182, 515 (1958). 



