Fluorescent stereomicroscope, microinjection training module and objective upgrade for C. elegans transgenic generation, husbandry and phenotypic screening

用于线虫转基因产生、饲养和表型筛选的荧光立体显微镜、显微注射训练模块和物镜升级

• 批准号: 458497-2014
• 负责人: Kormish, Jay
• 金额: $ 8.06万
• 依托单位: University of Manitoba
• 依托单位国家: 加拿大
• 项目类别: Research Tools and Instruments - Category 1 (<$150,000)
• 财政年份: 2013
• 资助国家: 加拿大
• 起止时间: 2013-01-01 至 2014-12-31
• 项目状态: 已结题
• 来源: https://www.nserc-crsng.gc.ca/ase-oro/Details-Detailles_eng.asp?id=541947
• 关键词: Fluorescent; stereomicroscope; microinjection; training; module

Organs are functional units of cells that perform specific tasks within an organism. An important question in the study of embryological development is how distinct cell types migrate, change shape and interact in a coordinated manner to form an organ. In my laboratory we use Caenorhabditis elegans, a free-living roundworm, with its powerful tools of genetics and live-imaging to study organ development. In particular, we study how the salivary gland cells of the worm's foregut use interactions with other cells of the foregut to form their elongated shape. Our research will combine multiple individual cellular interactions in space and time to form a 4-dimensional model of organ development. When compared to cell culture studies, this approach will provide a perspective of organ development that more closely models its true complexity. Small morphogenic changes in early organ development can impact final organ structure and, therefore, can provide a mechanism for the evolution of organ function and structure. The powerful tool of C. elegans genetics will identify the molecular pathways that are the driving force behind this change. The long-term goal of the research is to apply these basic findings to the development of the highly specialized foregut of the plant-parasitic nematodes, i.e., the agriculturally relevant Potato Cyst Nematode. Essential to my research program is the ability to generate transgenic animals. During the generation of transgenic nematodes, the DNA of genes required for salivary gland development is reintroduced into the animal. Modification of a gene's sequence is used to probe the function of that gene. C. elegans as a model organism also offers the power of genetic screens to identify mutations in important but uncharacterized genes regulating salivary gland development. Genetic screens and transgenic strain husbandry require a high-magnification fluorescent stereomicroscope. Defining changes in subcelluar structure require microscopes with high magnification objectives. The generation of transgenic animals requires a microinjection system. The requested equipment is essential for the advancement of my research program and the training of students in advanced microscopy and microtechniques.

器官是细胞的功能单位，在生物体内执行特定任务。胚胎发育研究中的一个重要问题是不同类型的细胞如何迁移，改变形状并以协调的方式相互作用以形成器官。在我的实验室里，我们使用秀丽隐杆线虫，一种自由生活的蛔虫，利用其强大的遗传学和实时成像工具来研究器官发育。特别是，我们研究了蠕虫的前肠的唾液腺细胞如何与前肠的其他细胞相互作用，形成其细长的形状。我们的研究将联合收割机结合多个个体细胞在空间和时间上的相互作用，形成器官发育的4维模型。与细胞培养研究相比，这种方法将提供一个器官发育的视角，更接近其真正的复杂性。早期器官发育中的微小形态发生变化可以影响最终器官结构，因此可以为器官功能和结构的进化提供机制。C. elegans遗传学将确定这一变化背后的驱动力的分子途径。这项研究的长期目标是将这些基本发现应用于植物寄生线虫高度特化的前肠的发育，即，与农业相关的马铃薯胞囊线虫。对我的研究计划来说，最重要的是培育转基因动物的能力。在转基因线虫的产生过程中，唾液腺发育所需基因的DNA被重新引入动物体内。基因序列的修饰用于探测该基因的功能。C.线虫作为模式生物也提供了遗传筛选的能力，以鉴定调节唾液腺发育的重要但未表征的基因的突变。遗传筛选和转基因菌株饲养需要高放大倍数的荧光体视显微镜。确定亚细胞结构的变化需要具有高放大倍数物镜的显微镜。转基因动物的产生需要显微注射系统。所要求的设备是必不可少的，我的研究计划的进步和学生在先进的显微镜和显微技术的培训。