Abstract
Similar to viruses, viroids can also be engineered and transformed into useful biotechnological tools. We describe here a viroid-based system to produce large amounts of recombinant RNA in Escherichia coli. A precursor of eggplant latent viroid (ELVd), with the RNA of interest inserted between positions U245 and U246, is co-expressed in E. coli along the chloroplastic isoform of the eggplant tRNA ligase, the enzyme that mediates the circularization of this viroid in the infected plants. In the bacterial cells, the chimeric ELVd-RNA-of-interest precursor self-cleaves through the embedded hammerhead ribozymes, and the monomer is recognized and circularized by the co-expressed tRNA ligase. The resulting circular RNA, likely bound to the tRNA ligase, accumulates to a high concentration in the bacterial cells.
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Acknowledgments
This work was supported by grants BIO2017-83184-R and BIO2017-91865-EXP from Ministerio de Ciencia e Innovación (Spain), co-financed by European Regional Development Fund (European Commission). BO is the recipient of a predoctoral contract from Universitat Politècnica de València (PAID-01-17).
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Ortolá, B., Daròs, JA. (2022). Production of Recombinant RNA in Escherichia coli Using Eggplant Latent Viroid as a Scaffold. In: Rao, A.L.N., Lavagi-Craddock, I., Vidalakis, G. (eds) Viroids. Methods in Molecular Biology, vol 2316. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-1464-8_25
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DOI: https://doi.org/10.1007/978-1-0716-1464-8_25
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