Core 2: In vivo cancer modeling and screening core facility

项目摘要/摘要：核心2 我们将建立一个用于体内癌症建模和筛查的核心设施。该核心设施(位于 癌症系统生物学中心)将由定义为模块的几组功能组件组成， 包括载体设计和基因组规模文库建设，大规模细胞培养和病毒生产， 肿瘤移植和体内联合筛选，CRISPR/Cas9介导的体细胞基因组编辑 侵袭性表型分析。在初步研究中，我们已经将体内体细胞基因组编辑应用于 在小鼠体内建立特定驱动基因的肿瘤模型，并在肺中进行体内筛选 转移动物模型。我们将在这个核心中构建四个模块，包括以下模块：模块1.设施 用于载体设计和基因组规模的文库构建。本模块支持以下功能调查 哺乳动物的基因组，可以揭示基因改变如何导致表型的改变，例如 项目1和2中描述的细胞侵袭性表型。模块2.大规模细胞培养的设施 培养、病毒载体生产和转导。这将用于两个目的：(1)产生 用于活体研究的定制病毒颗粒，涉及项目1和2以及其他项目的动物模型 合作者；(2)高通量筛选应用，需要转导具有 高度复杂的库。模块3.用于肿瘤移植和活体联合筛选的设备。这 模块将建立一个具有两个能力的设施：(1)肿瘤移植和(2)体内联合筛查。这 将用于验证在小鼠模型中的项目中发现的基因，以发现新的 使用体内筛选具有侵袭性表型的基因，或使合作研究能够发现新的 毒品目标。模块4.CRISPR/Cas9介导的体细胞基因组编辑工具，用于体细胞基因组 在各种动物中编辑小鼠基因组中的几乎任何一个基因座。我们将建立这个平台，并利用 它用于项目中描述的研究，并与耶鲁大学的各种研究人员建立合作。 具体地说，我们将利用CRISPR文库方法与细胞迁移的快速分析相结合 项目中描述的增强(RACE)系统，以确定基因改变如何导致 细胞侵袭性表型。此外，我们将利用核心中的这些模块来构建 WNK1、NKCC1及其靶点，以及突变的ERK、AKT和下游基因的组合。在……里面 综上所述，此核心将紧密集成到U54中心，提供强大的技术和资源 体内癌症系统生物学平台，并支持该中心以及其他研究项目 耶鲁大学和更广泛的科学界的合作者。因为耶鲁大学目前没有任何设施 提供类似的功能，这一新的核心将促进建立第一个体内癌症模型和 耶鲁大学的核心筛选设施。我们还将利用这一核心设施促进教育和培训，为 作为对癌症研究人员、科学家、学生和普通公众进行教育的基地。

PROJECT SUMMARY/ABSTRACT: Core 2 We will establish a core facility for in vivo cancer modeling and screening. This core facility (core 2 in the cancer systems biology center) will consist of several sets of functional components, defined as modules, including vector design and genome-scale library construction, large-scale cell culture and virus production, tumor transplantation and in vivo pooled screening, CRISPR/Cas9 mediated somatic genome editing for invasive phenotype analysis. In preliminary studies, we have applied in vivo somatic genome editing to generate tumor models of specific driver genes in mice, and demonstrated in vivo screening in a lung metastasis animal model. We will build four modules in this core, including the following: Module 1. Facility for vector design and genome-scale library construction. This module supports functional investigations of the mammalian genome that can reveal how genetic alterations lead to changes in phenotype, for example cellular invasiveness phenotype described in the Projects 1 and 2. Module 2. Facility for large-scale cell culture, viral vector production and transduction. This will serve for two purposes: (1) generation of customized viral particles for in vivo studies involving animals models for Projects 1 and 2 as well as other collaborators; (2) high-throughput screening applications that requires transduction of a population of cells with highly complex libraries. Module 3. Facility for tumor transplantation and in vivo pooled screening. This module will setup a facility with two capacities: (1) tumor transplantation and (2) in vivo pooled screening. This will serve for the purpose of validating the genes discovered in the projects in mouse models, to discover new genes with invasive phenotypes using in vivo screens, or to enable collaborative research for discovery of new drug targets. Module 4. Facility for CRISPR/Cas9 mediated somatic genome editing, for somatic genome editing in various animals for virtually any loci in the mouse genome. We will establish this platform and utilize it for research described in the projects and for setting up collaborations with various Yale investigators. Specifically, we will utilize CRISPR library approach in conjunction with the Rapid Analysis of Cell migration Enhancement (RACE) system described in the Projects, to identify how genetic alterations lead to changes in cellular invasiveness phenotype. In addition, we will utilize these modules in the core to build novel models of WNK1, NKCC1 and their targets, as well as combinations of mutant ERK, AKT and downstream genes. In summary, this core will be tightly integrated into the U54 center, provide a powerful technology and resource platform for in vivo cancer systems biology, and support Research Projects in the center as well as other collaborators at Yale and the wider scientific community. Since there is no facility currently existing at Yale providing similar functions, this new core will facilitate the establishment of the first in vivo cancer modeling and screening core facility at Yale. We will also utilize this core facility for promoting education and training, serving as a base for education of cancer researchers, scientists, students, and the general public.