Rescue of a plant cytorhabdovirus as versatile expression platforms for planthopper and cereal genomic studies
Rescue of a plant cytorhabdovirus as versatile expression platforms for planthopper and cereal genomic studies
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拯救植物细胞杆状病毒作为飞虱和谷物基因组研究的多功能表达平台
DOI：
10.1111/nph.15889
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发表时间：
2019-09-01
期刊：
影响因子：
9.4
通讯作者：
Wang, Xian-Bing
中科院分区：
文献类型：
--
作者：
Gao, Qiang;Xu, Wen-Ya;Wang, Xian-Bing
植物病毒已被用作基因组研究中用于异源蛋白表达的快速且成本有效的表达载体。然而，在单子叶植物和昆虫害虫中递送大的或多种外源蛋白是具有挑战性的。在这里，我们恢复了重组植物细胞弹状病毒，大麦黄纹花叶病毒（BYSMV），作为一个通用的表达平台，在谷物和小褐飞虱（SBPH，灰飞虱）昆虫载体。我们设计了BYSMV载体，为在大麦植物和SBPH中同时表达三种外源蛋白提供了通用的表达平台。此外，BYSMV载体能表达c. 600个氨基酸的β-葡萄糖醛酸酶（GUS）蛋白和红色荧光蛋白，其稳定地存在于谷物（包括小麦、大麦、谷子和玉米植物）的系统感染的叶和根中。此外，我们已经证明BYSMV载体可以用于大麦赤霉素（GA）生物合成基因的生物学功能的分析。在一项重大的技术进步中，BYSMV载体被开发用于同时递送CRISPR/Cas9核酸酶和单向导RNA，用于本氏烟草叶中的基因组编辑。综上所述，我们的研究结果提供了相当大的潜力，快速筛选功能蛋白质在谷物和稻飞虱，并开发其他昆虫传播的负链RNA病毒的有效方法。
Plant viruses have been used as rapid and cost-effective expression vectors for heterologous protein expression in genomic studies. However, delivering large or multiple foreign proteins in monocots and insect pests is challenging. Here, we recovered a recombinant plant cytorhabdovirus, Barley yellow striate mosaic virus (BYSMV), for use as a versatile expression platform in cereals and the small brown planthopper (SBPH, Laodelphax striatellus) insect vector. We engineered BYSMV vectors to provide versatile expression platforms for simultaneous expression of three foreign proteins in barley plants and SBPHs. Moreover, BYSMV vectors could express the c. 600-amino-acid beta-glucuronidase (GUS) protein and a red fluorescent protein stably in systemically infected leaves and roots of cereals, including wheat, barley, foxtail millet, and maize plants. Moreover, we have demonstrated that BYSMV vectors can be used in barley to analyze biological functions of gibberellic acid (GA) biosynthesis genes. In a major technical advance, BYSMV vectors were developed for simultaneous delivery of CRISPR/Cas9 nuclease and single guide RNAs for genomic editing in Nicotiana benthamiana leaves. Taken together, our results provide considerable potential for rapid screening of functional proteins in cereals and planthoppers, and an efficient approach for developing other insect-transmitted negative-strand RNA viruses.