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病毒诱导的基因沉默在植物基因功能研究中的应用及优势

Applications and advantages of virus-induced gene silencing for gene function studies in plants.

作者信息

Burch-Smith Tessa M, Anderson Jeffrey C, Martin Gregory B, Dinesh-Kumar S P

机构信息

Department of Molecular, Cellular, and Developmental Biology, Yale University, New Haven, CT 06520-8104, USA.

出版信息

Plant J. 2004 Sep;39(5):734-46. doi: 10.1111/j.1365-313X.2004.02158.x.

Abstract

Virus-induced gene silencing (VIGS) is a recently developed gene transcript suppression technique for characterizing the function of plant genes. The approach involves cloning a short sequence of a targeted plant gene into a viral delivery vector. The vector is used to infect a young plant, and in a few weeks natural defense mechanisms of the plant directed at suppressing virus replication also result in specific degradation of mRNAs from the endogenous plant gene that is targeted for silencing. VIGS is rapid (3-4 weeks from infection to silencing), does not require development of stable transformants, allows characterization of phenotypes that might be lethal in stable lines, and offers the potential to silence either individual or multiple members of a gene family. Here we briefly review the discoveries that led to the development of VIGS and what is known about the experimental requirements for effective silencing. We describe the methodology of VIGS and how it can be optimized and used for both forward and reverse genetics studies. Advantages and disadvantages of VIGS compared with other loss-of-function approaches available for plants are discussed, along with how the limitations of VIGS might be overcome. Examples are reviewed where VIGS has been used to provide important new insights into the roles of specific genes in plant development and plant defense responses. Finally, we examine the future prospects for VIGS as a powerful tool for assessing and characterizing the function of plant genes.

摘要

病毒诱导的基因沉默(VIGS)是一种最近开发的用于鉴定植物基因功能的基因转录抑制技术。该方法包括将目标植物基因的短序列克隆到病毒载体中。该载体用于感染幼苗,几周后,植物针对抑制病毒复制的天然防御机制也会导致来自靶向沉默的内源植物基因的mRNA发生特异性降解。VIGS速度快(从感染到沉默只需3 - 4周),不需要培育稳定的转化体,能够鉴定在稳定株系中可能致死的表型,并且有潜力沉默基因家族中的单个或多个成员。在这里,我们简要回顾导致VIGS发展的发现以及有效沉默的实验要求。我们描述了VIGS的方法及其如何优化并用于正向和反向遗传学研究。讨论了VIGS与其他植物功能缺失方法相比的优缺点,以及如何克服VIGS的局限性。回顾了VIGS用于为特定基因在植物发育和植物防御反应中的作用提供重要新见解的实例。最后,我们探讨了VIGS作为评估和鉴定植物基因功能的强大工具的未来前景。

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