REPLICATION OF POLIOVIRUS RNA STUDIED BY GEL FILTRATION AND ELECTROPHORESIS

The kinetics of synthesis of poliovirus RNA has been studied in suspension cultures of HeLa cells. Single stranded poliovirus RNA was separated from double stranded structures (replicative form and replicative intermediate) by gel filtration on sphere‐condensed agarose. Electrophoresis on polyacrylamide‐agarose gels was used for analytical separation of the three classes of poliovirus RNA in infected cells i. e. single stranded RNA, replicative form and replicative intermediate. Continuous labeling experiments confirmed that single stranded RNA is synthesized at a higher rate than the double stranded structures. Net synthesis of the replicative form levels off after 4 h of infection but the replicative form and the single stranded RNA are synthesized at high rates at later times. The replicative form probably precedes the single stranded RNA in the replication cycle. No free single strands complementary to viral RNA could be detected from 1 to 5 h after infection. Pulse labeling experiments reveal that the replicative form, the replicative intermediate and the single stranded RNA are synthesized at the highest rates 3–3.5 h post infection. The synthesis of the replicative intermediate and the single stranded RNA decreases at later times, while the replicative form synthesis continues unabated. At 3.5 h post infection a short (1 min) pulse introduces more label into the replicative intermediate than the single stranded RNA and the replicative form. As the length of the pulse increases the relative amount of label in the single stranded RNA increases, the replicative form remains constant and the replicative intermediate decreases. These results indicate a precursor‐product relationship between the replicative intermediate and the single stranded RNA, at 3.5 h post infection. At late times in the replication cycle most of the newly synthesized replicative form is not formed as an end product from the replicative intermediate. Copyright © 1969, Wiley Blackwell. All rights reserved