SGER: Developing Tools for the Analysis of Reproductive Barriers

高等植物中花粉和雌蕊之间种间屏障的分子性质目前尚不清楚。我们计划研究具有显著遗传和分子优势的重要作物品种番茄的种间屏障。然而，在我们开始对这些屏障进行分子分析之前，我们需要掌握一些工具。本提案的目标是开发这些工具。首先，我们将开发一个可视化系统，用于在雌蕊内生长花粉管。其次，我们将开发一种荧光原位杂交或“FISH-cassette”，用于染色体臂的标记和识别在生殖障碍中重要的连锁基因。该卡带将包含一个基于ds的工程转座元件，可用于诱变和增强子捕获。第三，我们将采用一项新兴技术（“分泌组”分析），并测试其对花粉和雌蕊分泌蛋白质检测的适用性。这些细胞和分子系统将在本项目之外的植物生物学研究中有广泛的应用。更广泛的影响这项工作将在几个层面上产生更广泛的影响。首先，科罗拉多州立大学的本科生将在整个项目期间参与工作的许多方面。如上所述，这项工作将影响植物研究界，因为所开发的工具将有广泛的应用。此外，这项工作将允许将基因组方法扩展到生物多样性研究，对驯化物种和野生物种进行比较。最后，植物繁殖的研究代表了一个对整个社会具有重要经济意义的领域，迄今为止，在基因组水平上理解繁殖的努力是有限的。对生殖障碍的认识与基因水平转移有直接关系；无论是在将理想的性状从野生物种转移到驯化物种方面，还是在限制转基因从作物转移到野生物种方面。项目outcomes1。拟南芥和番茄花粉启动子-绿色荧光蛋白融合将根据贝丁格实验室的要求在首次发表后提供给科学界。这些结构将对许多对花粉管生长和花粉-雌蕊相互作用感兴趣的研究人员有用。在资助期间产生的带有FISH磁带的最初转基因番茄品系将被扩充，并开始管理过程，为将来产生一系列完整的转基因品系做准备。一旦一套完整的转基因系（每条染色体臂上都有一个转基因盒）产生，这些将在茄科基因组学网络http://www.sgn.cornell.edu/上公布，并将提供给番茄研究界。来自分泌组研究的数据将存储在Jocelyn Rose博士正在开发的分泌组数据库中，该数据库可在他的网站http://labs.plantbio.cornell.edu/rose/上获得，此外，序列将存储在GenBank中。

Intellectual MeritThe molecular nature of interspecific barriers between pollen and pistils in higher plants is not currently understood. We plan to study interspecific barriers in tomato, an important crop species with significant genetic and molecular advantages. However, before we embark on the molecular analysis of these barriers, there are several tools we need to have in hand. The objective of this proposal is the development of these tools. First, we will develop a system for the visualization of growing pollen tubes within pistils. Second, we will develop a Fluorescent In Situ Hybridization or "FISH-cassette" for the marking of chromosome arms and identifying linked genes important in reproductive barriers. This cassette will contain a Ds-based engineered transposable element that can be used for both mutagenesis and enhancer trapping. Third, we will take an emerging technology ("secretome" analysis) and test its applicability to the detection of proteins secreted from pollen and pistils. These cellular and molecular systems will have a wide utility in plant biology research beyond this project.Broader ImpactsThe work will have a broader impact at several levels. First, undergraduate students at Colorado State University will be involved with many aspects of the work throughout the duration of the project. As noted above, the work will impact the plant research community, since the tools to be developed will have wide application. In addition, the work will permit the expansion of genomic approaches to biodiversity studies with the comparison of domesticated and wild species. Finally, research in plant reproduction represents an area of significant economic importance to society at large, and efforts toward understanding of reproduction at the genomic level have been limited to date. The understanding reproductive barriers is directly relevant to horizontal gene transfer; both in terms of transferring desirable traits from wild species into domesticated ones, and in terms of limiting transfer of transgenes from crops into wild species.Project outcomes1. Arabidopsis and tomato pollen promoter-GFP fusions will be available to the scientific community after initial publications upon request from the Bedinger lab. These constructs will be of use to numerous researchers with interests in pollen tube growth and pollen-pistil interactions.2. The initial transgenic tomato lines with a mapped FISH cassette generated during the funding period will be bulked up and the curation process initiated in preparation for generating a complete series of transgenic lines in the future. Once a complete set of transgenic lines (with a transgene cassette mapped to each chromosome arm) has been generated, these will be announced on the Solanaceae Genomics Network http://www.sgn.cornell.edu/ and will be made available to the tomato research community.3. Data from the secretome studies will be deposited in Dr. Jocelyn Rose's developing secretome database, available on his website http://labs.plantbio.cornell.edu/rose/ and sequences will additionally be deposited into GenBank.