Gunupuru Lokanadha R, Perochon Alexandre, Ali Shahin S, Scofield Steven R, Doohan Fiona M
Department of Plant, Food, and Environmental Sciences, Dalhousie University, Truro, NS, Canada.
School of Biology and Environmental Science and Earth Institute, College of Science, University College Dublin, Dublin, Ireland.
Methods Mol Biol. 2019;1900:95-114. doi: 10.1007/978-1-4939-8944-7_7.
With the recent advances in sequencing technologies, many studies are generating lists of candidate genes associated with specific traits. The major bottleneck in functional genomics is the validation of gene function. This is achieved by analyzing the effect of either gene silencing or overexpression on a specific phenotypic or biochemical trait. This usually requires the generation of stable transgenic plants and this can take considerable time. Therefore any technique that expedites the validation of gene function is of particular benefit in cereals, including barley. One such technique is Virus-Induced Gene Silencing (VIGS), which evokes a natural antiviral defense mechanism in plants. VIGS can be used to downregulate gene expression in a transient manner, but long enough to determine its effects on a specific phenotype. It is particularly useful for screening candidate genes and selecting those with potential for disease control. VIGS based on Barley Stripe Mosaic Virus (BSMV) is a powerful and efficient tool for the analysis of gene function in cereals. Here we present a BSMV VIGS protocol for simple and robust gene silencing in barley and describe it to evaluate the role of the hormone receptor BRI1 (Brassinosteroid Insensitive 1) in barley leaf resistance to Fusarium infection.
随着测序技术的最新进展，许多研究正在生成与特定性状相关的候选基因列表。功能基因组学的主要瓶颈是基因功能的验证。这是通过分析基因沉默或过表达对特定表型或生化性状的影响来实现的。这通常需要生成稳定的转基因植物，而这可能需要相当长的时间。因此，任何能够加快基因功能验证的技术对包括大麦在内的谷类作物都特别有益。一种这样的技术是病毒诱导基因沉默（VIGS），它能在植物中引发天然的抗病毒防御机制。VIGS可用于以瞬时方式下调基因表达，但时间足够长以确定其对特定表型的影响。它对于筛选候选基因和选择具有疾病控制潜力的基因特别有用。基于大麦条纹花叶病毒（BSMV）的VIGS是分析谷类作物基因功能的强大而有效的工具。在这里，我们展示了一种用于在大麦中进行简单而稳定的基因沉默的BSMV VIGS方案，并描述了它以评估激素受体BRI1（油菜素内酯不敏感1）在大麦叶片对镰刀菌感染的抗性中的作用。
