Expression of bacterial chloramphenicol acetyltransferase gene in tobacco plants mediated by TMV-RNA

Abstract

We have constructed three tobacco mosaic virus (TMV) cDNA derivatives by modification of the full-length cDNA clone from which infectious TMV-RNA can be transcribed in vitro. A coatless TMV construct lacks most of the coat protein gene and chimeric TMV constructs retain the bacterial chloramphenicol acetyltransferase (CAT) gene in place of the coat protein gene. When in vitro transcripts from these cDNA derivatives were inoculated on the local lesion tobacco plants, TMV-specific lesions were produced. In the case of the TMV-CAT chimeras, however, the lesions were small compared to those of wild-type TMV and those produced by transcript derived from the coatless construct. Northern blot analysis of RNA extracted from the inoculated leaves of the systemic host plants revealed replication of the derivative genomic RNAs and production of their own subgenomic RNAs corresponding to the coat protein mRNA. The TMV-CAT chimeras produced biologically active CAT in the inoculated leaves of the systemic host. CAT activity increased at least until 2 weeks post-inoculation and was approximately 0.1 units/mg of tissue at 10 days post-inoculation. Thus, TMV-RNA may be utilized as a new plant expression vector.