FIG. 3.

Rescue of synthetic DI-C RNA. With the plasmid described in Fig. 2, T7 transcripts with the information for DI-C RNA were obtained in vitro (A). The RNAs were transfected into ST cells that were previously infected with the helper virus (TGEV PUR46-MAD strain). Supernatants of the infected cultures were passaged six times (from P0 to P6), the RNAs of passages P4, P5, and P6 were extracted and analyzed by Northern hybridization with a probe complementary to the leader sequence (B). In lanes P4, P5, and P6, a band with the expected size of DI-C RNA was identified. The RNA from passage P6 of ST cells infected with the helper virus (HV) but untransfected was also analyzed. The numbers and letters on the left of the gel indicate number of the RNA and the protein that is encoded by this mRNA. S, spike; 3a, non-structural protein 3a; E, envelope protein; M, membrane protein; N, nucleoprotein.