Cytoskeletal Function in C. elegans Embryos

线虫胚胎中的细胞骨架功能

• 批准号: 10794146
• 负责人: BRUCE A BOWERMAN
• 金额: $ 1.1万
• 依托单位: UNIVERSITY OF OREGON
• 依托单位国家: 美国
• 财政年份: 2019
• 资助国家: 美国
• 起止时间: 2019-06-01 至 2024-11-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10794146
• 关键词: Administrative Supplement; Age; Animals; Biological Process; CRISPR/Cas technology; Caenorhabditis elegans; Cell division; Chimeric Proteins; Cytoskeleton; Embryo; Experimental Designs; Faculty; Funding; Gene Structure; Genes; Institution; Meiosis; Molecular Genetics; Oocytes; Research; Resolution; Structure; System; Technology; Transgenes; Transgenic Organisms; aged; genetic approach; genome editing; in vivo; light microscopy; member; mutant; optogenetics; protein function

We request funds to purchase two high capacity and versatile BioRad Thermal Cyclers. These will replace a single much older and lower capacity thermal cycler currently shared by my lab and the adjacent labs of my nearby colleagues and faculty members Chris Doe and Diana Libuda. Currently, due to the age of our machine and its limited capacity, and a lack of other institutional options, we have suffered from lack of access to amplification for constructing transgenic strains we need for our research. The discovery of CRISPR/Cas9 and its applications to genome editing have revolutionized molecular genetics in all systems, including C. elegans, and our limited thermal cycler capacity is now frequently limiting and slowing our progress. By purchasing two new higher capacity machines, and having a shared arrangement with two other labs, we will relieve not only the limitation to access in my own lab but also in the labs of two of my closest and physically adjacent colleagues. The remarkable advances in high resolution light microscopy make our ability to generate fluorescent protein fusions to endogenous loci more and more powerful, and we also are implementing more highly mechanistic structure/function studies that require the creation of transgenic lines that express mutant transgenes to replace endogenous genes. Finally, we also are now beginning to use optogenetics to more rigorously explore gene requirements. We need increased thermal cycler capacity to generate transgene constructs for use in all of these powerful molecular genetic approaches.

我们申请资金购买两台高容量和多功能的BioRad热循环仪。 这些将取代一个单一的旧得多和低容量的热循环仪目前 我的实验室和我附近的同事和教职员工的相邻实验室共享 克里斯·多伊和戴安娜·里瑟。目前，由于我们的机器的年龄和其有限的 能力，以及缺乏其他体制选择，我们因缺乏机会而遭受损失。 扩增以构建我们研究所需的转基因菌株。的 CRISPR/Cas9的发现及其在基因组编辑中的应用， 分子遗传学在所有系统，包括C。elegans和我们有限的热循环仪 能力现在常常限制和减缓我们的进展。通过购买两个新的 更高容量的机器，并与其他两个实验室共享，我们将 不仅可以在我自己的实验室里，而且可以在我的两个实验室里， 最亲密的和身体上相邻的同事。高分辨率的显著进步 光学显微镜使我们能够产生荧光蛋白融合到内源性 越来越强大，我们也在实施更高度的机械化， 结构/功能研究，需要建立转基因株系， 突变的转基因来取代内源基因。最后，我们现在也开始 利用光遗传学更严格地探索基因需求。我们需要增加 热循环仪产生转基因构建体的能力，用于所有这些强大的 分子遗传学方法