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Production of Circular Recombinant RNA in Escherichia coli Using Viroid Scaffolds.
Methods Mol Biol. 2021; 2323:99-107.MM

Abstract

Viroids are small circular, noncoding, highly base-paired RNAs able to infect higher plants. Recently, it has been shown that viroids can be used as very stable scaffolds to produce recombinant RNA in Escherichia coli. Coexpression of an RNA precursor consisting of a viroid monomer, in which the RNA of interest is inserted, flanked by domains of the viroid hammerhead ribozyme, along with a host plant tRNA ligase, the enzyme that catalyzes viroid circularization in infected plants, allows for accumulation of large amounts of the chimeric viroid-RNA of interest in E. coli. Since viroids do not replicate in E. coli, high accumulation most probably results from viroid scaffold stability, resistance to exonucleases due to circularity, and accumulation as a ribonucleoprotein complex with tRNA ligase. Purification of the recombinant RNA from total E. coli RNA is also facilitated by the circular structure of the product.

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Publisher Full Text (DOI)

Authors+Show Affiliations

Daròs JA
Instituto de Biología Molecular y Celular de Plantas (Consejo Superior de Investigaciones Científicas-Universitat Politècnica de València), Valencia, Spain. jadaros@ibmcp.upv.es.

MeSH

Cloning, MolecularDNA, ComplementaryEscherichia coliPlant ProteinsPlasmidsRNARNA Ligase (ATP)RNA, CatalyticRNA, CircularRNA, PlantRNA, ViralSolanum melongenaViroids

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

34086276

Citation

Daròs, José-Antonio. "Production of Circular Recombinant RNA in Escherichia Coli Using Viroid Scaffolds." Methods in Molecular Biology (Clifton, N.J.), vol. 2323, 2021, pp. 99-107.
Daròs JA. Production of Circular Recombinant RNA in Escherichia coli Using Viroid Scaffolds. Methods Mol Biol. 2021;2323:99-107.
Daròs, J. A. (2021). Production of Circular Recombinant RNA in Escherichia coli Using Viroid Scaffolds. Methods in Molecular Biology (Clifton, N.J.), 2323, 99-107. https://doi.org/10.1007/978-1-0716-1499-0_8
Daròs JA. Production of Circular Recombinant RNA in Escherichia Coli Using Viroid Scaffolds. Methods Mol Biol. 2021;2323:99-107. PubMed PMID: 34086276.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Production of Circular Recombinant RNA in Escherichia coli Using Viroid Scaffolds. A1 - Daròs,José-Antonio, PY - 2021/6/4/entrez PY - 2021/6/5/pubmed PY - 2021/8/20/medline KW - Circular RNA KW - Escherichia coli KW - Recombinant RNA KW - Viroid scaffold KW - tRNA ligase SP - 99 EP - 107 JF - Methods in molecular biology (Clifton, N.J.) JO - Methods Mol Biol VL - 2323 N2 - Viroids are small circular, noncoding, highly base-paired RNAs able to infect higher plants. Recently, it has been shown that viroids can be used as very stable scaffolds to produce recombinant RNA in Escherichia coli. Coexpression of an RNA precursor consisting of a viroid monomer, in which the RNA of interest is inserted, flanked by domains of the viroid hammerhead ribozyme, along with a host plant tRNA ligase, the enzyme that catalyzes viroid circularization in infected plants, allows for accumulation of large amounts of the chimeric viroid-RNA of interest in E. coli. Since viroids do not replicate in E. coli, high accumulation most probably results from viroid scaffold stability, resistance to exonucleases due to circularity, and accumulation as a ribonucleoprotein complex with tRNA ligase. Purification of the recombinant RNA from total E. coli RNA is also facilitated by the circular structure of the product. SN - 1940-6029 UR - https://www.unboundmedicine.com/medline/citation/34086276/Production_of_Circular_Recombinant_RNA_in_Escherichia_coli_Using_Viroid_Scaffolds_ DB - PRIME DP - Unbound Medicine ER -