quantitative microscopy-based rapid phenotyping and screening

描述(申请人提供)：突触是神经系统最基本的功能。线虫由于其基因组测序以及丰富的分子生物学工具和突变体，是寻找基因和阐明途径的优秀遗传模型系统。由于其神经系统的简单，线虫在了解神经系统模式和突触发育的分子机制方面已经取得了许多突破。然而，目前的瓶颈是手动和非定量技术，如视觉屏幕，通常既限制了实验的吞吐量，也限制了人们可以检查的表型。我们的长期目标是开发技术，以了解基因、年龄和环境如何共同定义并继续重塑有机体的神经系统。由于相关的长度尺度和自动化的可能性，微技术是线虫神经科学研究的理想选择；同样，定量成像技术是破译分子机制的关键。R01项目的目标是设计用于大规模实时成像和定量成像技术的微型设备，以研究体内系统中突触的发育。从这项研究中发现的基因和途径可能成为神经疾病治疗的靶点。我们假设，基于定量显微镜的方法确实能够识别传统方法无法识别的新基因和途径。该项目的第一个组成部分是开发芯片上快速和高含量的活体成像技术，同时开发用于分析此类高含量数据的算法和定量措施。该项目的第二个组成部分是使用这些新技术进行筛选和研究。这种方法是创新的，因为这里开发的技术极大地提高了现有成像和筛查工具的能力，速度提高了几个数量级，而且比传统的手动方法更灵敏和准确。这项拟议的研究意义重大，因为它满足了高通量和高内容筛选以及识别新基因和途径的迫切需求。此外，除了对特定神经生物学的贡献外，这些技术还广泛适用于发育细胞生物学等领域，以及其他小生物，如苍蝇幼虫和斑马鱼胚胎。

DESCRIPTION (provided by applicant): Synapses are most fundamental to the function of a nervous system. C. elegans is an excellent genetic model system for finding genes and elucidating pathways because of its sequenced genome and the abundance of molecular biology tools and mutants. Due to the simplicity of its nervous system, many breakthroughs have been made in C. elegans for understanding molecular mechanisms in the patterning of the nervous system and synapse development. The current bottlenecks, however, are in the manual and non-quantitative techniques such as visual screens, often limiting both the throughput of the experiments and the phenotypes one can examine. Our long-term objective is to develop technologies to understand how genes, age, and the environment together define and continue to remodel the nervous system of an organism. Microtechnologies are ideal for studies of C. elegans neuroscience because of the relevant length scales and the possibility for automation; similarly quantitative imaging techniques are key to deciphering molecular mechanisms. The objective of this R01 project is to engineer micro devices for large-scale live imaging and quantitative imaging technologies in order to study synapse development in an in vivo system. Genes and pathways emerging from this study could potentially become targets of therapeutics in neurological disorders. We hypothesize that quantitative microscopy-based approaches can indeed enable identification of novel genes and pathways that conventional approaches cannot. The first component of this project is to develop on-chip rapid and high-content in vivo imaging techniques, and in parallel to develop algorithms and quantitative measures for the analysis of such high-content data. The second component of the project is to perform screens and studies using these novel technologies. The approach is innovative because the technology developed here dramatically increases the capabilities of existing imaging and screening tools by several orders of magnitude in speed and much more sensitive and accurate than conventional manual approaches. The proposed research is significant because it fills the urgent need in high-throughput and high-content screens as well as identifying novel genes and pathways. In addition, besides the contribution to the specific neurobiology, the technologies are widely applicable to areas such as developmental cell biology, and to other small organisms such as fly larvae and zebrafish embryos.