268 FRAENKEL-CONRAT AND WILLIAMS 



thymus and RNA from turnip yellow A few preliminary experiments on 



mosaic virus ^^) yielded almost nucleic the properties of the reconstituted 



acid-free pellets which were inactive virus indicated somewhat greater labil- 



(Table 2). ity to alkali (pH 9) than that of TMV. 



Table 2 



Effect of Reaction Conditions on Regeneration of Virus Activity 



Reaction Conditions * 

 Protein (1 per cent) + RNA (0.1 per cent) : 



I minute, pH 6 

 24 hours 

 96 hours 



Protein (0.01 per cent) + RNA (0.001 per cent) : 



24 hours, pH 6 



96 hours 

 Protein + RNase-digested RNA 

 Protein + TYMV-RNA 

 Protein t 

 RNAt 



* Protein and RNA stand for the preparations isolated from TMV by the methods de- 

 scribed in the text. TYMV-RNA is the ribonucleic acid isolated from turnip yellow mosaic 

 virus. RNase is ribonuclease. 



t Reassayed dilute solution after one week. 



% Different preparations, assayed about 10 times, generally on separate plants (see n. 9) ; 

 in the customary half-leaf assay method, up to 0.5 lesions were occasionally obtained with the 

 same preparations. 



Ribonuclease (0.2 |_ig. for 5 mg.) also to assess the degree of homogeneity of 



appeared to decrease the activity. A the suspensions as well as to obtain 



fractionation of the nucleoprotein ma- counts and length distributions of the 



terial by centrifugation at 25,000 rpm. reconstituted rods. In the protein, the 



for 30 minutes yielded a pellet of con- only visible particles, aggregates pre- 



siderably higher infectivity than the sumably of the original 4.5 S material, 



residual half of the material, pelleted appeared as disks about 5-15 m^ thick 



at 40,000 rpm. (1 hour). Material so and with central holes. The diameter 



purified seemed to increase in activity of the disk was 15 m^ and that of the 



upon storage in 0.3-0.5 per cent solu- central hole about 4 m|j. (Fig. 2). 



tion at 3°. It also appeared to become The appearance of these perforated 



more resistant to alkali and to ribonu- disks is identical with that of partially 



clease. polymerized X-protein.^ In the nucleic 



Electron Microscopic Studies.— ¥Aec- acid solution only occasional poorly 

 tron microscopy was used qualitatively defined fibrils could be detected. The 

 to study the shapes and sizes of the reconstituted rods appeared to be iden- 

 components involved in the formation tical in shape and size with intact 

 of the virus rods and quantitatively ^^ TMV, except for a greater random- 

 ness of length (Fig. 3). 



10 Kindly supplied by Dr. S. S. Cohen and Preparations of the purified protein 



'^^"^YTrM."^^"^'"^' ^ r, , T^ ^ J and nucleic acid were examined for 



II Williams, R. C, Backus, R. C, and , . • -j r 

 Steere, R. L., /. Am. Chein. Soc. ly.lQSl, any electron microscopic evidence of 

 1951. contamination with intact TMV. In a 



