PDF(Pubmed)
Abstract：
In the post-genomic era, virus-induced gene silencing (VIGS) has played an important role in research on reverse genetics in plants. Commonly used Agrobacterium-mediated VIGS inoculation methods include stem scratching, leaf infiltration, use of agrodrench, and air-brush spraying. In this study, we developed a root wounding-immersion method in which 1/3 of the plant root (length) was cut and immersed in a tobacco rattle virus (TRV)1:TRV2 mixed solution for 30 min. We optimized the procedure in Nicotiana benthamiana and successfully silenced N. benthamiana, tomato (Solanum lycopersicum), pepper (Capsicum annuum L.), eggplant (Solanum melongena), and Arabidopsis thaliana phytoene desaturase (PDS), and we observed the movement of green fluorescent protein (GFP) from the roots to the stem and leaves. The silencing rate of PDS in N. benthamiana and tomato was 95-100%. In addition, we successfully silenced two disease-resistance genes, SITL5 and SITL6, to decrease disease resistance in tomatoes (CLN2037E). The root wounding-immersion method can be used to inoculate large batches of plants in a short time and with high efficiency, and fresh bacterial infusions can be reused several times. The most important aspect of the root wounding-immersion method is its application to plant species susceptible to root inoculation, as well as its ability to inoculate seedlings from early growth stages. This method offers a means to conduct large-scale functional genome screening in plants.
摘要：
在后基因组时代,病毒诱导基因沉默(VIGS)在植物反向遗传学研究中发挥了重要作用。常用的农杆菌介导的VIGS接种方法包括茎刮，叶入渗,使用农田，和喷枪喷涂。在这项研究中,我们开发了一种根损伤-浸入方法，其中将植物根的1/3（长度）切开并浸入烟草摇铃病毒（TRV）1：TRV2混合溶液中30分钟。我们优化了Nicotianabenthamiana的程序，并成功地使N.benthamiana沉默，番茄（番茄），辣椒（辣椒），茄子（茄子），和拟南芥八烯去饱和酶（PDS），我们观察到绿色荧光蛋白（GFP）从根部到茎和叶的运动。PDS在N.benthamiana和番茄中的沉默率为95-100％。此外,我们成功地沉默了两个抗病基因,SITL5和SITL6，下降番茄的抗病性（CLN2037E）。根伤浸法可在短时间内,高效率地大批量接种植物,新鲜的细菌输液可以重复使用几次。根伤浸泡法的最重要方面是将其应用于对根接种敏感的植物物种，以及它从早期生长阶段接种幼苗的能力。该方法提供了在植物中进行大规模功能基因组筛选的手段。
