Replication of Semliki Forest virus RNA in infected chick cells

Doctoral thesis English OPEN
Martin, Brian Anthony Bernard
  • Subject: QR355

1. RNA synthesis was studied in chick cells infected with SFV, to determine the sizes and the structures of the RNA species made, and the mechanism of replication of SFV RNA.\ud \ud 2. Four single-stranded (SS), three double-stranded (DS) and multi-stranded (MS) species of RNA were detected in infected cells. Methods were described for their separation..\ud \ud 3. The molecular weights of the SS species of RNA were 4.0 x 10 6 , 3.1 x 10 6 , 2.3 x 10 6 and 1.8 x 10 6 , the largest of which was virion RNA. The molecular weights of the three DS RNA species, determined by separation of the strands of the DS RNA and analysis of the product, 6 6 6 were 8.0 x 10 , 5.6 x 10 , and 3.6 x 10 •\ud 4. Viral RNA synthesis was detected within ½ hour after infection and was logarithmic for about 3 hours. The rate of RNA synthesis was linear for the next 5 hours, and then decreased rapidly. \ud \ud 5. DS and MS RNA were the first viral species detectable, at 1 hour after infection, No SS RNA was detected up to 11/2 hours after infection when 42S RNA was detected.\ud \ud 6. During the linear phase of RNA synthesis, all species of RNA could be detected after labelling infected cells for 1 minute. The ratios of labelling of the SS species of RNA were the same for pulses varying between 1 minute and 70 minutes. More label was detected in DS and MS RNA in pulses up to 2 minutes, but :in pulses of 5 minutes or more, the SS RNA species were more highly labelled.\ud \ud 7. No alteration of the pattern of viral RNA species was seen when RNA was extracted :in the presence of DEPC, suggesting that no major alterations in the structure of RNA were taking p:).&ce during extraction. It was concluded that DS and MS RNA represent viral RF and RI, respectively.\ud \ud 8. The synthetic times for 26s and 425 RNA molecules were estimated to be 58 seconds and 130 seconds, respectively.\ud \ud 9. The evidence suggests that the mechanism of SFV RNA replication is largely semi-conservative. A model for the replication of SFV RNA is described, which involves the synthesis of all the species of SS RNA anemone template, corresponding :in length to 42S RNA.\ud \ud 10. No SS RNA complementary to virion RNA was detected in infected cells, suggesting that viral messenger RNA was of the same polarity as virion RNA.\ud \ud 11. Thus, SFV appears to belong to animal virus class IV, although it shares some features with class V viruses.\ud
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