ShRNA Screening and Histopathology

ShRNA 筛选和组织病理学

• 批准号: 280454646
• 负责人: Professor Dr. Martin Eilers
• 金额: --
• 依托单位: Pathologisches Institut
• 依托单位国家: 德国
• 项目类别: Research Units
• 财政年份: 2015
• 资助国家: 德国
• 起止时间: 2014-12-31 至 2022-12-31
• 项目状态: 已结题
• 来源: https://gepris.dfg.de/gepris/projekt/280454646?language=en
• 关键词: ShRNA; Screening; Histopathology

This central project provides critical technologies to this research consortium. Specifically, Z01 has three tasks:First, this project provides the synthesis of focused shRNA libraries and and the analysis of screens with these libraries. In particular, the project provides focused shRNA libraries as well as technical advice for both in vitro and in vivo shRNA screens, advises individual projects on set-up of in vivo experiments (e.g. animal number), and recovers and sequences the genomic DNA containing the retroviral inserts. The project also analyses the sequencing data including a statistical analysis. As part of this service, the Z01 also handles all communication with Mirimus and the handling of shRNA-transgenic mice that are imported into the consortium. The second task of this project is the target validation using a pipeline of established stress assays. This part involves the analysis of multiple stress parameters after depletion of specific targets or interference with small molecules using high-content microscopy and optical image analysis. These assays are established and data are provided as quantitative evaluation to all members of this consortium. This screening core facility will also perform small molecule screens of FDA-approved compounds in the next funding period. The third task of this project is to provide mouse and human pathology analyses with the aim to validate therapeutic targets (e.g. by analyzing their expression in human tumors) and to study the in vivo effects of interfering with essential processes in tumors. In the next funding period, this part of the project will include a standardized analysis of immune cell infiltrates into tumors after therapeutic intervention. This analysis pipeline and the necessary staining protocols have been established.

该中心项目为该研究联盟提供关键技术。具体来说，Z01有三个任务：首先，本项目提供重点shRNA文库的合成，并利用这些文库进行筛选分析。特别是，该项目提供集中的shRNA文库以及体外和体内shRNA筛选的技术建议，就体内实验的设置（例如动物数量）向个别项目提供建议，并恢复含有逆转录病毒插入物的基因组DNA并对其进行测序。该项目还分析了测序数据，包括统计分析。作为这项服务的一部分，Z01还处理与Mirimus的所有通信，以及对导入该联盟的shrna转基因小鼠的处理。该项目的第二个任务是使用已建立的应力分析管道进行目标验证。这部分涉及使用高含量显微镜和光学图像分析分析特定靶点耗尽或小分子干扰后的多个应力参数。建立了这些检测方法，并将数据作为定量评价提供给该联盟的所有成员。该筛选核心设施还将在下一个资助期内对fda批准的化合物进行小分子筛选。该项目的第三个任务是提供小鼠和人类病理分析，目的是验证治疗靶点（例如，通过分析它们在人类肿瘤中的表达），并研究干扰肿瘤基本过程的体内效应。在下一个资助期，该项目的这一部分将包括治疗干预后免疫细胞浸润肿瘤的标准化分析。该分析管道和必要的染色方案已经建立。