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https://hdl.handle.net/20.500.12955/1803
Título : | Simplifying plant gene silencing and genome editing logistics by a one-Agrobacterium system for simultaneous delivery of multipartite virus vectors |
Autor : | Aragonés, Verónica Aliaga, Flavio Pasin, Fabio Daròs, José Antonio |
Fecha de publicación : | 14-jul-2022 |
Resumen : | Viral vectors provide a quick and effective way to express exogenous sequences in eukaryotic cells and to engineer eukaryotic genomes through the delivery of CRISPR/Cas components. Here, we present JoinTRV, an improved vector system based on tobacco rattle virus (TRV) that simplifies gene silencing and genome editing logistics. Our system consists of two mini T-DNA vectors from which TRV RNA1 (pLX-TRV1) and an engineered version of TRV RNA2 (pLX-TRV2) are expressed. The two vectors have compatible origins that allow their cotransformation and maintenance into a single Agrobacterium cell, as well as their simultaneous delivery to plants by a one-Agrobacterium/two-vector approach. The JoinTRV vectors are substantially smaller than those of any known TRV vector system, and pLX-TRV2 can be easily customized to express desired sequences by one-step digestion-ligation and homology-based cloning. The system was successfully used in Nicotiana benthamiana for launching TRV infection, for recombinant protein production, as well as for robust virus-induced gene silencing (VIGS) of endogenous transcripts using bacterial suspensions at low optical densities. JoinTRV-mediated delivery of single-guide RNAs in a Cas9 transgenic host allowed somatic cell editing efficiencies of ≈90%; editing events were heritable and >50% of the progeny seedlings showed mutations at the targeted loci. |
Palabras clave : | CRISPR/Cas9 Heritable gene editing PLX binary vector multiplexing Tobacco rattle virus Virus-induced gene silencing (VIGS) Virus-induced genome editing (VIGE) |
Editorial : | Wiley |
Citación : | Aragonés, V.; Aliaga, F.; Pasin, F. & Darós, J. (2022). Simplifying plant gene silencing and genome editing logistics by a one-Agrobacterium system for simultaneous delivery of multipartite virus vectors. Biotechnology Journal Volume 17, Issue 7, 2100504. doi: 10.1002/biot.202100504 |
URI : | https://hdl.handle.net/20.500.12955/1803 |
metadata.dc.identifier.doi: | https://doi.org/10.1002/biot.202100504 |
metadata.dc.subject.ocde: | https://purl.org/pe-repo/ocde/ford#4.04.00 |
Aparece en las colecciones: | Artículos científicos |
Ficheros en este ítem:
Fichero | Descripción | Tamaño | Formato | |
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Aragonés-et-al_2022_VIGS_VIGE.pdf | 7,87 MB | Adobe PDF | Visualizar/Abrir |
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