Cell line Core

细胞系核心

基本信息

批准号：
10202526
负责人：
Gaorav P Gupta
金额：
$ 12.48万
依托单位：
UNIV OF NORTH CAROLINA CHAPEL HILL
依托单位国家：
美国
项目类别：
财政年份：
2020
资助国家：
美国
起止时间：
2020-07-01 至 2026-06-30
项目状态：
未结题

来源：
https://reporter.nih.gov/project-details/10202526
关键词：
Aliquot Alleles Biological Biological Assay CRISPR/Cas technology Cell Line Cells Clone Cells Clustered Regularly Interspaced Short Palindromic Repeats Collaborations Communities Credentialing Cryopreservation Custom DNA Double Strand Break DNA Repair Enzymes DNA-Directed DNA Polymerase Doxycycline Engineering Ensure Enzymes Experimental Designs Future Gene Targeting Generations Genes Genetic Engineering Genome Genome engineering Genomic Instability Genomics Genotype Goals Human In Vitro Karyotype Karyotype determination procedure Knock-in Knock-out Ligands Malignant Neoplasms Mammalian Cell Manuscripts Mediating Methods Modeling Monitor Mus Mycoplasma Polymerase Process Proteins Protocols documentation Publications Quality Control Reagent Reproducibility Research Research Activity Research Personnel Role Secure Services Standardization Structure of thyroid parafollicular cell System Testing Time Transgenes Validation base cost effectiveness design digital empowered experimental study genetically modified cells homologous recombination in vivo interest laboratory experiment multidisciplinary overexpression parity programs protein expression quality assurance repaired repository screening service programs success synergism transcriptome sequencing transgene delivery transgene expression vector

Aliquot Alleles Biological Biological Assay CRISPR/Cas technology Cell Line Cells Clone Cells Clustered Regularly Interspaced Short Palindromic Repeats Collaborations Communities Credentialing Cryopreservation Custom DNA Double Strand Break DNA Repair Enzymes DNA-Directed DNA Polymerase Doxycycline Engineering Ensure Enzymes Experimental Designs Future Gene Targeting Generations Genes Genetic Engineering Genome Genome engineering Genomic Instability Genomics Genotype Goals Human In Vitro Karyotype Karyotype determination procedure Knock-in Knock-out Ligands Malignant Neoplasms Mammalian Cell Manuscripts Mediating Methods Modeling Monitor Mus Mycoplasma Polymerase Process Proteins Protocols documentation Publications Quality Control Reagent Reproducibility Research Research Activity Research Personnel Role Secure Services Standardization Structure of thyroid parafollicular cell System Testing Time Transgenes Validation base cost effectiveness design digital empowered experimental study genetically modified cells homologous recombination in vivo interest laboratory experiment multidisciplinary overexpression parity programs protein expression quality assurance repaired repository screening service programs success synergism transcriptome sequencing transgene delivery transgene expression vector

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项目摘要

PROJECT SUMMARY This project will investigate mammalian DNA polymerase θ (Pol θ), the defining enzyme for repair of DNA double-strand breaks by polymerase theta-mediated end joining (TMEJ). This is Core C (“Cell Line Engineering Core”) which is part of a Program Project titled, “Polymerase theta, genome instability, and cancer”. Core C will be responsible for providing native and genetically engineered cell lines to support all of the cell-based assays performed in Projects 1, 2, and 4 of the Program Project. The authentication of mammalian cell line reagents is vital for rigor and reproducibility of laboratory experiments. By establishing and implementing standardized protocols for cell line validation, efficient genome manipulation, cryopreservation, and dissemination, the Core will ensure quality control of cell line reagents and minimize the likelihood of erroneous observations and interpretations. In Aim 1, “Credentialing of mammalian cell line models utilized in this Program Project”, we will implement a standardized and comprehensive protocol for authentication and characterization of mammalian cell line models used across the Program Project. In Aim 2, “CRISPR-based cell line genome engineering services for the Program Project”, we will provide expert services in the design and execution of customized cell line genome engineering tasks in support of Program Project goals. Cas9 protein utilized in this Aim will be provided by Core B. In Aim 3, “Controlled and stable expression of transgenes in mammalian cell lines”, we will utilize doxycycline-inducible and constitutive transposon vectors for the generation of cell lines that express a variety of mutagenized and/or tagged transgenes of interest. Core C services will be highly coordinated within the Program Project, and will facilitate rigor and reproducibility, as well as cost-effectiveness, of highly integrated research activities aimed at understanding the role and targetability of DNA Polymerase q in cancer.

项目摘要该项目将研究哺乳动物DNA聚合酶θ（polθ），定义酶修复DNA 通过聚合酶theta介导的末端连接（TMEJ）的双链断裂。这是核心C（“单元线工程核心”）是一个计划项目的一部分，标题为“聚合酶theta，基因组不稳定性和癌症”。核心C将负责提供本地和一般的工程细胞系，以支持所有在项目项目的项目1、2和4中执行的基于单元的阿萨斯。哺乳动物细胞系试剂的身份验证对于严格和实验室的可重复性至关重要实验。通过建立和实施用于细胞系验证的标准化协议，有效的基因组操纵，冷冻保存和传播，核心将确保细胞系试剂的质量控制和最小化错误的观察和解释的可能性。在AIM 1中，“该程序项目中使用的哺乳动物细胞系模型的证书”，我们将实施哺乳动物细胞系的身份验证和表征的标准化和全面的协议整个程序项目中使用的模型。在AIM 2中，“基于CRISPR的细胞系基因组工程服务为程序项目”，我们将提供定制细胞系基因组工程任务的设计和执行专家服务以支持计划项目目标。 CAS9在此目标中使用的CAS9蛋白将由CoreB提供。在AIM 3中，“哺乳动物细胞系中翻译的受控和稳定表达”，我们将利用强力霉素诱导和本构的转座子向量，用于生成表达多样的细胞系感兴趣的诱变和/或标记的翻译。核心C服务将在计划项目中高度协调，并将促进严格和高度集成研究活动的可重复性以及成本效益，旨在了解 DNA聚合酶Q在癌症中的作用和目标性。