Recombinant simian virus 40 (SV40) virus genomes have been constructed in vitro by joining SVGT1, the segment of SV40 DNA between map co-ordinates 0.15 and 0.73 (clockwise), to either the Escherichia coli gene for thymidine kinase ( Ecotdk), or one of the Saccharomyces cerevisae genes for tRNA Tyr ( ScetyrG); the resulting recombinants were propagated in CV-1 monkey cells at 41 °C using tsA58 as a helper. The sEcotdk gene was obtained first as an approximately 20 kb ‡ DNA segment in a defective transducing phage, φ80 dtdk5, then as overlapping 1.85 and 2.35 kb segments in pMB9- Ecotdk, prior to insertion into SVGT1. The ScetyrG segment introduced into SVGT1 was a 1.25 kb DNA fragment contained in λgtl- ScetyrG, a recombinant that had been cloned previously by Olson et al. (1979). After infection of CV-1 monkey cells with the SVGT1- Ecotdk or SVGT1- ScetyrG hybrid viruses, RNA complementary to the exogenous DNA was produced. With SVGT1- Ecotdk the RNA homologous to the Ecotdk segment was heterogeneous, ranging in size from 1 to > 8 kb in length. At least 30 to 40% of this RNA was covalently joined to SV40-specific RNA. No E. coli thymidine kinase enzyme activity could be detected in the infected cells. By contrast, infection with SVTG 1- ScetyrG resulted in the formation of a transfer RNA-sized RNA, complementary to the tRNA Tyr coding sequence of the ScetyrG segment, as well as a population of heterogenous large RNAs. Since the formation of the tRNA-sized RNA occurred after infection with recombinants having the ScetyrG segments in either of the two alternative orientations in SVGT1, it is possible that transcription of the tyrG sequence is initiated within the cloned segment. ‡ Abbreviation used: kb, kilobase (10 3 bases)
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