Functional Genomics of Maize Gametophytes

PI：Matthew Evans(卡内基华盛顿州立大学)Copis：Donald Auger(南达科他州立大学；子获奖者)，John Fowler和Scott Givan(俄勒冈州立大学，子获奖者)，Erik Vollbrecht(爱荷华州立大学，子获奖者)合作者：Kelly Beck，Gabriel Garcia(斯坦福大学，子获奖者)了解配子体功能的遗传基础将对农业产生重要影响，因为配子体对植物繁殖至关重要。雌、雄配子体组成被子植物生活史的单倍体阶段，在减数分裂之后立即形成种子(胚胎和胚乳)。虽然配子体很小，细胞分裂很少，但它们对生殖至关重要，因为它们产生配子，控制受精过程，并影响种子的发育。然而，由于对这些单倍体组织有害的突变很难恢复和维持，因此对配子体功能和发育的遗传和细胞机制知之甚少，特别是在农作物中。该项目试图使用模式作物玉米(玉米)特有的遗传工具来克服这一限制，以完成配子体所需基因的基因组规模的研究。通过配子体传递重复染色体区域的三体种群将用于筛选激活子转座子标记的配子体致死突变体。这些突变体的表型将在雄配子体和雌配子体中进行表征，并确定相应的突变基因的DNA序列。将使用多个等位基因和RNA表达分析来验证选定的一组候选基因的身份。在配子体表达谱和生物信息学中的互补目标--识别其他植物模型中的同源基因并整合该项目产生的数据--将允许评估配子体功能的遗传基础。最后，将使用RNA干扰来测试跨物种边界预测配子体功能的能力，以在双子叶模型拟南芥中针对选定的基因。更广泛的影响：该项目与寻求影响植物繁殖的许多农业目标有关--例如，控制杂交种子生产的花粉育性，限制花粉介导的转基因流动，以及诱导无融合生殖。由于创建的工具和种群以及生成的序列将免费提供给科学界，该项目将使其他研究人员能够通过创建配子体特定的序列索引突变集合来更好地研究配子体。该项目创建的所有库存都将存放在玉米鸡舍库存中心(maizecoop.cropori.uiuc.edu)。所有ac转座子侧翼序列都可以通过BLAST进行搜索，既可以在PlantGDB(www.PLANGDB.org)上搜索，也可以在S数据库：ZGamDB(maizegametophyte.org)上搜索。表型和表达数据也将在ZGamDb上访问。为了长期存储和广泛的社区访问，表型和遗传数据将被纳入MaizeGDB(www.maizegdb.org)，表达数据将存储在基因表达总览(www.ncbi.nlm.nih.gov/geo)和植物表达数据库(www.plexdb.org)。该项目将帮助培训10名本科生，让他们接触植物科学研究，成为K-12学生的科学导师。与斯坦福大学S哈斯公共服务中心合作，这些本科生导师将被安置在人口众多的社区，这些群体来自代表性不足的群体，并帮助K-12学生发展科学体验。此外，该项目将培训两名博士后研究人员和一名研究生在基因组规模的植物生物学方法方面，并将向本科生和高中生介绍基因组科学。

PI: Matthew Evans (Carnegie Institution of Washington)CoPIs: Donald Auger (South Dakota State University; subawardee), John Fowler and Scott Givan (Oregon State University; subawardees), Erik Vollbrecht (Iowa State University; subawardee)Collaborator: Kelly Beck, Gabriel Garcia (Stanford University; subawardees)An understanding of the genetic basis of gametophyte function will have important implications for agriculture, as gametophytes are central to plant reproduction. The female and male gametophytes make up the haploid phase of the angiosperm life cycle, which immediately succeeds meiosis and precedes formation of the seed (embryo and endosperm). Although gametophytes are small and undergo few cell divisions, they are crucial for reproduction, as they produce gametes, control the fertilization process, and influence development of the seed. However, because mutations that are deleterious to these haploid tissues are difficult to recover and maintain, relatively little is known about the genetic and cellular mechanisms underlying gametophyte function and development, especially in crop plants. This project seeks to overcome this limitation using genetic tools unique to the model crop Zea mays (maize) to accomplish a genomic-scale investigation of gametophytically-required genes. Trisomic stocks that transmit duplicate chromosomal regions through the gametophyte will be used to screen for Activator transposon-tagged gametophyte-lethal mutants. The phenotype of these mutants will be characterized in male and female gametophytes, and the corresponding DNA sequences of the mutated genes identified. Verification of the identity of a select group of candidate genes will be accomplished using multiple alleles and RNA expression analyses. Complementary aims in expression profiling of gametophytes and in bioinformatics - to identify orthologous genes in other plant models and integrate the data generated by the project - will allow an assessment of the genetic basis of gametophyte function. Finally, the ability to predict gametophytic functions across species boundaries will be tested using RNA interference to target select genes in the dicot model Arabidopsis thaliana. Broader Impacts:This project is relevant to many agricultural objectives seeking to influence plant reproduction - for example, controlling pollen fertility for hybrid seed production, limiting pollen-mediated transgene flow, and inducing apomixis. Because the tools and stocks created, and sequences generated, will be freely available to the scientific community, the project will enable other researchers to better investigate gametophytes through creation of a gametophyte-specific sequence-indexed mutant collection. All stocks created by the project will be deposited in the Maize Coop Stock Center (maizecoop.cropsci.uiuc.edu). All Ac transposon flanking sequences will be searchable by BLAST, both at PlantGDB (www.plantgdb.org), and at the project''s database: ZGamDB (maizegametophyte.org). Phenotypic and expression data will also be accessible at ZGamDb. For long-term storage and broad community access, phenotypic and genetic data will be incorporated into MaizeGDB (www.maizegdb.org), and expression data will be deposited at the Gene Expression Omnibus (www.ncbi.nlm.nih.gov/geo) and the Plant Expression Database (www.plexdb.org).The project will help train 10 undergraduates to serve as science mentors for K-12 students, through exposing them to plant science research. In partnership with Stanford University''s Haas Center for Public Service, these undergraduate mentors will be placed in communities with large populations from under-represented groups, and help develop science experiences for K-12 students. In addition, the project will train two postdoctoral researchers and a graduate student in genomic-scale approaches to plant biology, and will introduce undergraduates and high school students to genomic science.