Institutes of Green Bio Science and Technology, Seoul National University, Pyeongchang 25354, Republic of Korea.
Horticultural and Herbal Crop Environment Division, National Institute of Horticultural and Herbal Science, Wanju 55365, Republic of Korea.
Plant Physiol. 2019 Nov;181(3):867-880. doi: 10.1104/pp.19.00836. Epub 2019 Sep 3.
While pepper () is a highly recalcitrant species for genetic transformation studies, plant virus-based vectors can provide alternative and powerful tools for transient regulation and functional analysis of genes of interest in pepper. In this study, we established an effective virus-based vector system applicable for transient gain- and loss-of-function studies in pepper using (BBWV2). We engineered BBWV2 as a dual gene expression vector for simultaneous expression of two recombinant proteins in pepper cells. In addition, we established enhanced and stable expression of recombinant proteins from the BBWV2-based dual vector via coexpression of a heterologous viral suppressor of RNA silencing. We also developed a BBWV2-based virus-induced gene silencing (VIGS) vector, and we successfully silenced the phytoene desaturase gene () using the BBWV2-based VIGS vector in various pepper cultivars. Additionally, we optimized the BBWV2-based VIGS system in pepper by testing the efficiency of gene silencing under different conditions. This BBWV2-based vector system represents a convenient approach for rapid and simple analysis of gene functions in pepper.
辣椒是一种遗传转化研究中高度抗逆的物种，而基于植物病毒的载体可以为辣椒中感兴趣基因的瞬时调控和功能分析提供替代的、强大的工具。在本研究中，我们利用 （BBWV2）建立了一个有效的基于病毒的载体系统，可用于辣椒中的瞬时功能获得和功能丧失研究。我们将 BBWV2 设计为一个双基因表达载体，可在辣椒细胞中同时表达两种重组蛋白。此外，我们通过共表达一种异源病毒 RNA 沉默抑制子，实现了基于 BBWV2 的双载体中重组蛋白的增强和稳定表达。我们还开发了一种基于 BBWV2 的病毒诱导基因沉默（VIGS）载体，并成功地使用基于 BBWV2 的 VIGS 载体在各种辣椒品种中沉默了类胡萝卜素脱饱和酶基因（）。此外，我们通过在不同条件下测试基因沉默的效率，对基于 BBWV2 的 VIGS 系统在辣椒中的优化进行了测试。该基于 BBWV2 的载体系统为快速简便地分析辣椒中的基因功能提供了一种便捷的方法。
