Genome-Wide Hunt For Metal Hyperaccumulation Genes

[12977447] david E. salt .该项目的总体目标是在金属超富集植物中鉴定与金属超富集有关的基因。这些独特的植物物种能够以Cd、Ni、Se或Zn的形式积累0.1%至3%的茎干生物量，具体取决于物种。这些特性在目前利用植物清洁金属污染场地（植物修复）的努力中可能是无价的。已知的金属超蓄积体由于体积小、生长缓慢，不适合植物修复。然而，它们是金属超积累基因的独特来源。超过25%的已知超蓄积物物种是芸苔科的成员，因此它们与拟南芥有关。通过研究拟南芥相关物种金属超积累的分子遗传学，研究者将利用在拟南芥基因组计划中开发的技术和遗传资源，利用强大的功能基因组学技术在遗传水平上剖析金属超积累。将收集来自世界各地的芸苔科金属超富集物，并利用三种互补的方法鉴定与超富集有关的重要基因。将收集来自北美、法国、德国、奥地利、意大利、希腊和土耳其的芸苔科20多种超积累植物的近40个品种的种子。发现适合T-DNA插入突变的金属超蓄积体的加入将被识别，并将产生超过100,000个遗传系。在正向遗传方法中，这些细胞系将筛选表现出金属敏感性和金属超积累损失的突变体。将从这些突变体中分离出T-DNA标记基因，并确定其在金属超积累中的作用。在逆向遗传方法中，基因组DNA池将从这些品系中产生，并通过PCR筛选，以识别已知或怀疑与金属超积累有关的基因中含有T-DNA插入的品系。拟南芥基因组序列将为这种反向遗传方法提供丰富的候选基因来源。在第三种方法中，cDNA表达文库将在大肠杆菌和酵母菌中建立，并筛选赋予重金属抗性和敏感性的基因。综上所述，这些方法将为鉴定与植物金属超积累有关的基因提供一个全面的框架。该项目所鉴定的一组基因将为未来开发适合金属污染场地植物修复的植物提供宝贵资源。该项目由美国环境保护局、国家科学基金会、海军研究办公室和战略环境研究与发展计划共同发起的植物修复联合计划资助。

01297747David E. SaltThe overall objective of this project is to identify genes involved in metal hyperaccumulation in metal-hyperaccumulating plants. These unique plant species are able to accumulate between 0.1 and 3% of their shoot dry biomass as Cd, Ni, Se or Zn depending on the species. Such characteristics could be invaluable in current efforts to use plants to clean metal-contaminated sites (phytoremediation). The known metal hyperaccumulators are not well suited for phytoremediation because of their small size and slow growth. However, they are a unique source of genes for metal hyperaccumulation. Over 25% of the known hyperaccumulator species are members of the Brassicaceae family, and as such they are related to Arabidopsis thaliana. By investigating the molecular genetics of metal hyperaccumulation in species related to A. thaliana, the investigators will utilize the technical and genetic resources developed during the Arabidopsis genome project, harnessing powerful functional genomics technologies to dissect metal hyperaccumulation at the genetic level. Metal hyperaccumulators in the Brassicaceae will be collected from around the world, and genes important in hyperaccumulation will be identified using three complementary approaches. Seed from approximately 40 accessions of over 20 different species of hyperaccumulators in the Brassicaceae family will be collected from North America, France, Germany, Austria, Italy, Greece and Turkey. Accessions of metal hyperaccumulators found to be amenable to T-DNA insertional mutagenesis will be identified and over 100,000 genetic lines will be generated. In a forward genetic approach, these lines will be screened for mutants exhibiting metal-sensitivity and loss of metal-hyperaccumulation. T-DNA tagged genes from these mutants will be isolated, and their role in metal hyperaccumulation determined. In a reverse genetic approach, genomic DNA pools will be generated from these lines and screened by PCR to identify lines containing T-DNA insertions in genes already known or suspected to be involved in metal hyperaccumulation. The A. thaliana genome sequence will provide a rich source of candidate genes for this reverse genetic approach. In a third approach, cDNA expression libraries will be created in E. coli and yeast from hyperaccumulating species and screened for genes conferring heavy-metal resistance and sensitivity. Taken together these approaches will provide a comprehensive framework for the identification of genes involved in metal hyperaccumulation in plants. The set of genes identified in this project will provide a valuable resource for the future development of plants ideally suited for the phytoremediation of metal polluted sites. This project was funded through the Joint Program on Phytoremediation, co-sponsored by the Environmental Protection Agency, the National Science Foundation, the Office of Naval Research, and the Strategic Environmental Research and Development Program.