In vitro transcription of infectious RNAs from full-length cDNAs of tobacco mosaic virus - PubMed (original) (raw)

In vitro transcription of infectious RNAs from full-length cDNAs of tobacco mosaic virus

T Meshi et al. Proc Natl Acad Sci U S A. 1986 Jul.

Abstract

We have cloned full-length double-stranded cDNAs of tobacco mosaic virus (TMV) (tomato strain L) RNA into a transcription vector, pPM1, which facilitates the correct transcription initiation from the first nucleotide of the inserted double-stranded cDNA, corresponding to the 5' end of TMV RNA. When plasmid DNA is linearized at a unique restriction site (Mlu I) introduced just downstream of the double-stranded cDNA insert and used as a template for in vitro transcription by Escherichia coli RNA polymerase in the presence of m(7)GpppG, the transcribed RNAs are infectious for tobacco plants. A simple reconstitution procedure increases the infectivity >100 times. Unexpectedly, both the uncapped transcript and the transcript from the uncut plasmid DNA are also infectious, although their infectivities are very low. The progeny viruses multiplying in tobacco plants accurately reflect the cloned sequence. By the same method, we succeeded in the in vitro transcription of infectious RNA of attenuated strain L(11)A, which is phenotypically distinguishable from wild-type TMV on both tobacco and tomato plants.

PubMed Disclaimer

References

    1. J Mol Biol. 1980 Apr;138(2):179-207 - PubMed
    1. Adv Virus Res. 1981;26:145-99 - PubMed
    1. Proc Natl Acad Sci U S A. 1984 Nov;81(22):7066-70 - PubMed
    1. Nucleic Acids Res. 1975 Jul;2(7):1189-201 - PubMed
    1. FEBS Lett. 1976 Aug 15;67(2):209-13 - PubMed

LinkOut - more resources