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一种基于植物弹状病毒的多功能载体,用于基因沉默、miRNA表达与缺失以及抗体生产。

A Versatile Plant Rhabdovirus-Based Vector for Gene Silencing, miRNA Expression and Depletion, and Antibody Production.

作者信息

Peng Xingxing, Ma Xiaonan, Lu Shuting, Li Zhenghe

机构信息

State Key Laboratory of Rice Biology, Institute of Biotechnology, Zhejiang University, Hangzhou, China.

Ministry of Agriculture Key Laboratory of Molecular Biology of Crop Pathogens and Insect Pests, Zhejiang University, Hangzhou, China.

出版信息

Front Plant Sci. 2021 Jan 12;11:627880. doi: 10.3389/fpls.2020.627880. eCollection 2020.

DOI:10.3389/fpls.2020.627880
PMID:33510764
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC7835261/
Abstract

Plant virus vectors are ideal tools for delivery of genetic cargo into host cells for functional genomics studies and protein overexpression. Although a vast number of plant virus vectors have been developed for different purposes, the utility of a particular virus vector is generally limited. Here, we report a multipurpose plant rhabdovirus-based vector system suitable for a wide range of applications in . We engineered sonchus yellow net rhabdovirus (SYNV)-based gene silencing vectors through expressing a sense, antisense, or double-stranded RNAs of target genes. Robust target gene silencing was also achieved with an SYNV vector expressing a designed artificial microRNA. In addition, ectopic expression of a short tandem target mimic RNA using the SYNV vector led to a significant depletion of the target miR165/166 and caused abnormal leaf development. More importantly, SYNV was able to harbor two expression cassettes that permitted simultaneous RNA silencing and overexpression of large reporter gene. This dual capacity vector also enabled systemic expression of a whole-molecule monoclonal antibody consisting of light and heavy chains. These results highlight the utility of the SYNV vector system in gene function studies and agricultural biotechnology and provide a technical template for developing similar vectors of other economically important plant rhabdoviruses.

摘要

植物病毒载体是将遗传物质导入宿主细胞以进行功能基因组学研究和蛋白质过表达的理想工具。尽管已经开发了大量用于不同目的的植物病毒载体,但特定病毒载体的实用性通常是有限的。在此,我们报道了一种基于弹状病毒的多功能植物载体系统,适用于多种应用。我们通过表达靶基因的正义、反义或双链RNA,构建了基于南芥黄花叶病毒(SYNV)的基因沉默载体。使用表达设计的人工微小RNA的SYNV载体也实现了强大的靶基因沉默。此外,使用SYNV载体异位表达短串联靶标模拟RNA导致靶标miR165/166显著减少,并导致叶片发育异常。更重要的是,SYNV能够容纳两个表达盒,允许同时进行RNA沉默和大型报告基因的过表达。这种双功能载体还能够全身表达由轻链和重链组成的全分子单克隆抗体。这些结果突出了SYNV载体系统在基因功能研究和农业生物技术中的实用性,并为开发其他具有经济重要性的植物弹状病毒的类似载体提供了技术模板。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b34c/7835261/e4ef9559c789/fpls-11-627880-g006.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b34c/7835261/b5a327c41d0a/fpls-11-627880-g001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b34c/7835261/a0228ad29c6c/fpls-11-627880-g002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b34c/7835261/5c0cd0acf3e7/fpls-11-627880-g003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b34c/7835261/097459e0774d/fpls-11-627880-g004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b34c/7835261/e4615b30c067/fpls-11-627880-g005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b34c/7835261/e4ef9559c789/fpls-11-627880-g006.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b34c/7835261/b5a327c41d0a/fpls-11-627880-g001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b34c/7835261/a0228ad29c6c/fpls-11-627880-g002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b34c/7835261/5c0cd0acf3e7/fpls-11-627880-g003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b34c/7835261/097459e0774d/fpls-11-627880-g004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b34c/7835261/e4615b30c067/fpls-11-627880-g005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b34c/7835261/e4ef9559c789/fpls-11-627880-g006.jpg

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