CU Boulder Center for Cryo-ET (CCET)

科罗拉多大学博尔德分校冷冻电子断层扫描中心 (CCET)

• 批准号: 10261985
• 负责人: ANDREAS HOENGER
• 金额: $ 78.06万
• 依托单位: UNIVERSITY OF COLORADO
• 依托单位国家: 美国
• 财政年份: 2020
• 资助国家: 美国
• 起止时间: 2020-09-08 至 2024-08-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/10261985
• 关键词: Award; Biochemistry; Budgets; Cells; Cellular Structures; Collaborations; Colorado; Communities; Complex; Computer software; Cryo-electron tomography; Cryoelectron Microscopy; Crystallography; Data Collection; Educational workshop; Electron Microscopy; Electrons; Equipment; Funding; Home environment; Image Analysis; Instruction; International; Ions; Methods; Microscope; Names; Nucleic Acids; Organelles; Organization and Administration; Preparation; Procedures; Process; Proteins; Resources; Roentgen Rays; Running; Scientist; Services; Specimen; Synchrotrons; Technology; Tissues; Titan; Training; computerized tools; cost; detector; macromolecular assembly; macromolecule; software development; training opportunity

All funds awarded for Yr-01 are anticipated to be expended as planned and not available for re- budgeting for the equipment costs detailed in the budget. We will establish a satellite service center within a larger network according to RFAs RM-19-009 and -010, named the CU-Boulder Center for Cryo-Electron Tomography (CCET). Our satellite will focus on cryospecimen preparation and cryo-electron tomography (cryo-ET) applied to cells, organelles and large supramolecular assemblies. The applicants combine the expertise of three P.I.s, Michael Stowell (highresolution cryo-electron microscopy (cryo-EM)), Karolin Luger (biochemistry of large complex assemblies, especially protein-nucleic acid interactions), and Andreas Hoenger (former P41 director, cryo-EM and cryo-ET from macromolecules to cells and tissues). With our current staff, we combine extensive expertise with cryo-ET and relevant software. We will offer a broad range of cryo-electron microscopy (cryo-EM) and cryo-ET preparation procedures for cellular structures and intracellular macromolecular assemblies, as well as training opportunities for scientists who would like to establish these technologies, or at least part of them, for their home labs. Here in Boulder, we have several high-end microscopes available, including a newly installed Titan-Krios. Three of our microscopes are equipped with direct electron detectors. In addition, we have a comprehensive periphery for cryo-specimen preparation, storage, and handling resources, including computational tools. We will add cryo- Focused Ion Beam (cryo-FIB) milling to produce vitrified lamellae of cells and tissues. We propose the following organization of the facility: Specific Aim 1: Cryo-ET specimen preparation methods offered for service: Our technology and expertise will be made accessible for our users and for further collaborations with the data collection hub proposed in a parallel RFA (RM-19-010). Specific Aim 2: Cross-training opportunities forservice users: We will offer comprehensive training for users, including one-on-one instruction on microscopes and related equipment. As appropriate, we will pool users into small groups and run a training week as a hands-on workshop on equipment as well as related software developed by our facility such as Serial-EM, IMOD and PEET. While the emphasis with training will be on practical aspects of specimen preparation and data collection, we often encounter novice users in strong need for a thorough introduction into image analysis software as well. Specific Aim 3: Administration and organization of the facility: Users, assigned to us via the network-wide application process will have the opportunity to participate in the specimen refinement process, and receive training on the cryo-ET processes involved. Projects with high potential will be referred for high-throughput data collection to the central hub facility of the network. Boulder, Colorado is centrally located and well-connected nationwide through Denver International Airport.

预计所有YR-01授予的所有资金将被按计划支出，无法重新使用 预算中详细介绍的设备费用的预算。 我们将根据RFAS RM-19-009在较大的网络中建立一个卫星服务中心 和-010，名为Cu-Boulder Cryo-Electron断层扫描中心（CCET）。我们的 卫星将专注于施用的冰冻膜制剂和冷冻电子层析成像（Cryo-Et） 到细胞，细胞器和大分子组件。申请人结合了专业知识 迈克尔·斯托维尔（Michael Stowell Luger（大型复合组件的生物化学，尤其是蛋白质核酸相互作用）， 和Andreas Hoenger（前P41导演，Cryo-EM和Cryo-Et从大分子到细胞 和组织）。与当前的员工一起，我们将广泛的专业知识与冷冻-ET和相关性相结合 软件。我们将提供广泛的低温电子显微镜（Cryo-EM）和Cryo-ET 细胞结构和细胞内大分子组件的制备程序，AS 以及希望建立这些技术或在的科学家的培训机会 其中至少是他们的家庭实验室。在博尔德，我们有几个高端显微镜 可用，包括新安装的Titan-Krios。我们的三个显微镜配备了 直接电子探测器。此外，我们还有一个针对冷冻特异性的全面的外围 准备，存储和处理资源，包括计算工具。我们将添加cryo- 聚焦离子束（冷冻纤维）铣削以产生细胞和组织的玻璃薄片。我们 提出以下设施的组织： 特定目的1：提供服务的冷冻标本准备方法：我们的 技术和专业知识将可以为我们的用户访问并进行进一步的合作 在平行RFA（RM-19-010）中提出的数据收集中心。 特定目的2：跨培训机会提供服务用户：我们将提供全面的 对用户的培训，包括有关显微镜和相关设备的一对一指导。作为 合适的是，我们将成为小组的用户，并作为动手进行培训周 设备和相关软件的研讨会，我们的设施（例如串行EM）， IMOD和PEET。 虽然培训的重点将放在标本准备和数据的实际方面 收集，我们经常遇到新手用户，非常需要彻底介绍 图像分析软件。 具体目标 3：设施的管理和组织：用户，通过网络范围内分配给我们 申请过程将有机会参与标本完善过程， 并接受有关涉及的冷冻过程的培训。潜力很高的项目将是 将高通量数据收集转介给网络的中央集线器设施。博尔德， 科罗拉多州位于市中心，在全国范围内通过丹佛建立了良好的联系 国际机场。