SGER: Development of Tandem RNAi for Silencing of Gene Families in Rice

PI：G.埃里克·斯皮尔，达特茅斯学院CoPI：约瑟夫·J·基伯（北卡罗来纳大学教堂山分校）和帕梅拉·罗纳德（加州大学戴维斯分校） 分析植物基因作用的一个主要挑战是发现许多基因存在于基因家族中。由于基因家族成员之间的功能重叠（遗传冗余），因此必须破坏多个家族成员以确定其功能并改变其在植物中的作用。为了克服遗传冗余，我们提出了一种新的方法来同时破坏植物中的多个基因，通过靶向一个家族中多个基因的RNAi串联阵列来实现这一点。本项目将在单子叶水稻中开发和测试这种“超级沉默”系统。为此目的，将制备载体的组成型和诱导型版本。参与植物激素细胞分裂素和乙烯信号传导的三个基因家族将成为破坏的目标。将使用分子和生理学方法表征含有超沉默构建体的转基因系，以评估该方法的成功。更广泛的影响：超级沉默方法的成功将导致建立一种新的遗传工具，用于研究和修改水稻和其他单子叶作物物种的遗传控制性状。这种方法将能够快速修饰有助于改善农艺重要性状的基因和途径，如产量、非生物和生物抗性、胁迫反应和生物量。用于超级沉默的载体将提供给植物研究界。拟议的研究将通过为项目本科生和研究生提供实践培训，加强研究和教育的基础设施。

PI: G. Eric Schaller, Dartmouth College CoPIs: Joseph J. Kieber (University of North Carolina-Chapel Hill) and Pamela Ronald, (University of California, Davis) A major challenge to analyzing the roles of genes in plants has been the discovery that many genes are present in gene families. Due to functional overlap (genetic redundancy) among the members of a gene family, multiple family members must thus be disrupted in order to ascertain their function and to modify their role in the plant. To overcome genetic redundancy, we propose a novel approach to disrupt multiple genes simultaneously within a plant, accomplishing this by means of a tandem array of RNAi targeted to multiple genes within a family. This project will develop and test this "supersilencing" system in the monocot rice. For this purpose, both constitutive and inducible versions of the vectors will be made. Three gene families involved in signaling by the plant hormones cytokinin and ethylene will be targeted for disruption. Transgenic lines containing the supersilencing constructs will be characterized using molecular and physiological methods to assess the success of the approach. Broader impacts: The success of the supersilencing approach would result in the establishment of a new genetic tool for the study and modification of genetically controlled traits in rice and other monocot crop species. This approach would enable rapid modification of genes and pathways instrumental to the improvement of traits of agronomic importance such as yield, abiotic and biotic resistance, stress responses, and biomass. Vectors for use in supersilencing will be made available to the plant research community. The proposed research will enhance the infrastructure of research and education by providing hands-on training for project undergraduate students and graduate students.