Sequencing and Bioinformatics

测序和生物信息学

• 批准号: 10508449
• 负责人: Andrew Laurence Routh
• 金额: $ 21.62万
• 依托单位: SEATTLE CHILDREN'S HOSPITAL
• 依托单位国家: 美国
• 财政年份: 2022
• 资助国家: 美国
• 起止时间: 2022-06-22 至 2027-03-31
• 项目状态: 未结题
• 来源: https://reporter.nih.gov/project-details/10508449
• 关键词: Area; Behavior; Bioinformatics; Biological; Biology; Capsid; Cell Culture Techniques; Cells; Chromatin Structure; Chromosome Structures; Clinical; Collaborations; Communities; Complementary DNA; Computer Analysis; Computer software; Core Facility; DNA Sequencing Facility; Data; Databases; Development; Drug resistance; Environment; Event; Evolution; Experimental Designs; Exposure to; Gene Expression; Genetic Recombination; Genome; Genomic approach; HIV; HIV Genome; Human Resources; In Situ; In Vitro; Individual; Integrase; Length; Location; Measures; Messenger RNA; Minority; Molecular; Molecular Biology; Molecular Virology; Mutation; Nucleic Acids; Nucleotides; Nuts; Output; Pattern; Pharmacotherapy; Polymerase; Process; Protein Isoforms; Proteins; RNA; RNA Splicing; RNA Viruses; RNA-Protein Interaction; Research; Research Personnel; Research Project Grants; Resolution; Resources; Role; Sampling; Services; Single Nucleotide Polymorphism; Site; Specimen; Structure; System; Techniques; Technology; Training; Universities; Variant; Viral; Viral Genome; Virus; Virus Replication; Work; antiretroviral therapy; base; chromosomal location; crosslink; experience; experimental study; genome sequencing; innovation; insight; integration site; molecular sequence database; nanopore; new technology; next generation; next generation sequencing; novel; novel strategies; particle; pressure; resistance mutation; response; sequencing platform; single cell sequencing; single cell technology; single molecule; skills; small molecule inhibitor; tool; transcriptome sequencing; transcriptomics; viral genomics; virology

ABSTRACT, CORE 4 Next-Generation Sequencing is an essential and fundamental approach in molecular virology. In vitro, it can capture and characterize protein-nucleic interactions and RNA structure. In cell culture and in clinical settings, it can measure rates of viral polymerase error, viral response to small molecule inhibitors or changes in host gene expression. NGS can chart viral evolution, patterns of genome conservation, characterize origins of drug resistance and discover new viruses. As such, NGS has been transformative in our ability to study virus diversity and evolution and is still a burgeoning field. The Sequencing Core comprises a group of experts with experience with the implementation of the wide range of approaches in terms of molecular biology, experimental design, and downstream computational analyses. We will provide expert support and resources to provide Next- Generation Sequencing and downstream computational analyses to the B-HIVE Research Projects. Importantly, a key focus of the core will be to develop and apply approaches that go beyond what is accessible via commercial and university core facilities. We will provide and develop state-of-the-art technologies in close collaboration with the Research Projects that will help provide unique insights into HIV biology and evolution, whilst also providing novel platforms for deployments in other viral systems. A central focus of the B-HIVE is to understand the molecular mechanisms governing HIV entry, integration, particle assembly, and egress. Each of these processes involve profiling the viral and host nucleic acids during the viral lifecycle. The sequencing core will support the entire B-HIVE reflecting stages throughout the viral replication cycle. We will develop single-cell sequencing approaches, leveraging our extensive expertise in this area, to characterize the effects of integration site location, chromatin structure, HIV mutations, and exposure to latency reversing agents (LRAs) on proviral expression. We will apply and advance long-read sequencing technologies using the Oxford Nanopore Technologies MinION platform for both host transcriptomics and full- length HIV genome sequencing to characterize host and virus splicing. We will develop and implement novel approaches in virus genomics to characterize virus sequence diversity in both clinical and experimental settings in support of the main Research Projects. The Sequencing Core has extensive experience developing, deploying and maintaining new computational software and sequencing databases. These will provide an important resource to the B-HIVE centers, as well as allow the rapid dissemination and sharing of capabilities with the broader HIV and virus research community. Finally, we will provide training in each of our respective platforms and technologies in view of disseminating these skills throughout the B-HIVE and to support the development and progression of trainees working with the B-HIVE and its collaborators. 1

摘要，核心4 下一代测序技术是分子病毒学研究中的一个重要和基础的方法。在体外，它可以 捕获和表征蛋白质-核酸相互作用和RNA结构。在细胞培养和临床环境中， 可以测量病毒聚合酶错误率、病毒对小分子抑制剂的反应或宿主基因的变化 表情NGS可以绘制病毒进化，基因组保守模式，表征药物来源， 并发现新的病毒。因此，NGS在我们研究病毒多样性的能力方面具有变革性 和进化，仍然是一个新兴的领域。测序核心由一组经验丰富的专家组成 随着分子生物学，实验设计， 和下游计算分析。我们将提供专家支持和资源，以提供下一个- B-HIVE研究项目的世代测序和下游计算分析。 重要的是，核心的一个关键重点将是制定和应用超越可获得的方法 通过商业和大学核心设施。我们将提供和开发最先进的技术， 与研究项目的合作，这将有助于提供对艾滋病毒生物学和进化的独特见解， 同时还提供用于在其它病毒系统中部署的新平台。 B-HIVE的一个中心焦点是了解控制HIV进入、整合 粒子集合和出口这些过程中的每一个都涉及在病毒感染期间对病毒和宿主核酸进行分析。 病毒的生命周期测序核心将支持整个B-HIVE，反映整个病毒的各个阶段。 复制周期我们将开发单细胞测序方法，利用我们在这方面的广泛专业知识， 区域，以表征整合位点位置、染色质结构、HIV突变和暴露于 潜伏期逆转剂（LRA）对前病毒表达的影响。我们将应用并推进长读序测序 技术使用Oxford Nanopore Technologies MinION平台进行宿主转录组学和全基因组学研究， 长度HIV基因组测序以表征宿主和病毒剪接。我们将开发和实施新的 在临床和实验环境中表征病毒序列多样性的病毒基因组学方法 支持主要研究项目。Sequencing Core拥有丰富的开发、部署 并维护新的计算软件和测序数据库。这将提供一个重要的 资源的B-HIVE中心，以及允许快速传播和共享的能力， 更广泛的艾滋病毒和病毒研究界。最后，我们将在各自的平台上提供培训 为了在整个B-HIVE中传播这些技能，并支持开发 以及与B-HIVE及其合作者一起工作的学员的进步。 1