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Development of a novel activation tagging system that facilitates functional assignment of plant genes

开发一种新型激活标签系统，促进植物基因的功能分配

基本信息

批准号：
18510171
负责人：
NAKAJIMA Keiji
金额：
$ 2.61万
依托单位：
Nara Institute of Science and Technology
依托单位国家：
日本
项目类别：
Grant-in-Aid for Scientific Research (C)
财政年份：
2006
资助国家：
日本
起止时间：
2006 至 2007
项目状态：
已结题

来源：
https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-18510171/
关键词：
Arabidopsis Gene Activation tagging transposon transcription factor アクティべーションタギング

项目摘要

This research project aims to establish a "second generation" CAL4/UAS activation tagging system by improving the previous one. The new system requires three plant transformation constructs; (1) tissue-specific promoter:GVG, (2) Heat-shock promoter:Ac transposase and (3) Ds:UAS tagging vector Constructs (1) and (2) were introduced into wild-type Arabidosis thaliana plants and several homozygous transformants had been established for each construct. These two plant lines were genetically crossed and plant lines double homozygous for the two constructs were selected. These plants were then used as a host strain in the new tagging system. The host plants were transformed with the DsUAS tagging vector by floral dip method. Seeds thus obtained were then selected by Basta-resistance. Transformation efficiency was estimated to be more than 1%, indicating that 1000 transformants required to cover the entire Arabidopsis genome can be obtained by dipping as few as 20 host plants. In the future, it is required to test if the heat-shock promoter-Ac can transpose DsUAS DNA fragment in the genome of starter lines as designed initially. The basic materials for the "second generation" GAL4/CAS activation tagging system had been established by this project and it can be utilized to screen for novel mutants.

本研究旨在通过对原系统的改进，建立“第二代”CAL 4/UAS活化标记系统。新系统需要三种植物转化构建体：（1）组织特异性启动子：GVG，（2）热激启动子：Ac转座酶和（3）Ds：UAS标记载体将构建体（1）和（2）引入野生型拟南芥植物中，并且已经为每个构建体建立了几个纯合转化体。将这两个植物系进行遗传杂交，并选择对于两个构建体双纯合的植物系。然后将这些植物用作新标记系统中的宿主菌株。通过浸花法将DsUAS标签载体转化到寄主植物上。然后通过Basta抗性选择由此获得的种子。转化效率估计超过1%，表明覆盖整个拟南芥基因组所需的1000个转化体可以通过浸渍少至20个宿主植物来获得。热激启动子Ac能否按设计将DsUAS DNA片段转座到起始系基因组中，有待进一步研究。本研究为“第二代”GAL 4/CAS激活标签系统的构建奠定了基础，可用于筛选新的突变体。