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. 2005 Jun;79(11):7024–7041. doi: 10.1128/JVI.79.11.7024-7041.2005

FIG. 8.

FIG. 8.

CVB3/TD strains package negative-strand RNA. (A) Dilutions of T7 RNA polymerase-transcribed positive-strand RNA and (B) T3 RNA polymerase-transcribed negative-strand RNA from a linearized CVB3 subclone were slot blotted. (C) RNA was prepared from virus stocks pretreated with RNase as described in Materials and Methods using 104, 103 and 102 TCID50 units of CVB3/28 (CVB3 10,000, 1,000, and 100, respectively) or 107 rTCID50 TD8, 107 rTCID50 TD13, 105 rTCID50 TD18, 108 rTCID50 TD31, or 108 rTCID50 TD50. Blots were probed with 32P-labeled E1 (positive strand) or 5Puff (negative strand). (D) Strand-specific RNAs (+, positive; -, negative) were purified from T7 and T3 RNA polymerase transcripts of a subclone of CVB3/28 cDNA, mixtures of transcripts, and RNA of CsCl gradient-purified CVB3/28, CVB3/TD8, and CVB3/TD50 as described in Materials and Methods. RNA (0.3 μg) was used for each preparation. cDNAs prepared from these RNAs were amplified with KS1 and KS2, electrophoresed, and visualized as described in Materials and Methods. Lane M, 100-bp ladder. The arrow indicates a 600-bp band.