A Versatile Set of Ligation-Independent Cloning Vectors for Functional Studies in Plants
A Versatile Set of Ligation-Independent Cloning Vectors for Functional Studies in Plants
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DOI：
10.1104/pp.111.177337
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发表时间：
2011-07-01
期刊：
影响因子：
7.4
通讯作者：
Weijers, Dolf
中科院分区：
文献类型：
--
作者：
De Rybel, Bert;van den Berg, Willy;Weijers, Dolf
随着植物分子生物学进入组学时代，需要简单的克隆策略，允许高通量系统地研究大量基因的表达和功能。这样的策略将有助于分析基因（子）家族和/或转录组学鉴定的共表达基因集。在这里，我们提供了一套34连接非依赖性克隆（LIC）的二元载体的表达分析，蛋白质定位研究，和错误表达，将免费提供。这套植物LIC载体提供了一个快速的替代标准克隆策略，涉及连接酶或重组酶技术。我们展示了使用这种策略和我们的新载体，通过分析属于基本螺旋-环-螺旋转录因子家族的两个亚支的基因的表达结构域。我们发现，无论是最接近的同系物的目标monopteros 7（TMO 7/ATBS 1），也没有成员的ATBS 1相互作用因子亚支的假定TMO 7相互作用在胚胎中表达，并有非常有限的共表达在初级根分生组织。这表明这些基本的螺旋-环-螺旋转录因子很可能不参与TMO 7依赖的根分生组织的起始。
With plant molecular biology entering the omics era, there is a need for simple cloning strategies that allow high throughput to systematically study the expression and function of large numbers of genes. Such strategies would facilitate the analysis of gene (sub) families and/or sets of coexpressed genes identified by transcriptomics. Here, we provide a set of 34 ligation-independent cloning (LIC) binary vectors for expression analysis, protein localization studies, and misexpression that will be made freely available. This set of plant LIC vectors offers a fast alternative to standard cloning strategies involving ligase or recombination enzyme technology. We demonstrate the use of this strategy and our new vectors by analyzing the expression domains of genes belonging to two subclades of the basic helix-loop-helix transcription factor family. We show that neither the closest homologs of TARGET OF MONOPTEROS7 (TMO7/ATBS1) nor the members of the ATBS1 INTERACTING FACTOR subclade of putative TMO7 interactors are expressed in the embryo and that there is very limited coexpression in the primary root meristem. This suggests that these basic helix-loop-helix transcription factors are most likely not involved in TMO7-dependent root meristem initiation.