Expression Profiling of Cellular Metabolism Using Massively Parallel Sequencing

描述(由申请人提供)： 下一代测序(NGS)提供了一种强大的新技术，能够以简单、全面和经济的方式快速有效地询问DNA序列。使用NGS探测全基因组表达模式，定义转录因子靶向的全部基因，并评估全基因组甲基化状态已变得可行和负担得起。这笔赠款的目标是建立一个高吞吐量的NGS设施，以支持一批有实施新技术历史的科学家正在进行的研究项目，这些科学家有实施新技术的历史，以做出直接应用于人类健康的高影响的基础发现。这项研究的重点是导致死亡和残疾的两个主要原因：胆固醇代谢紊乱和燃料稳态失调。需要资金从应用生物系统公司购买一套寡核苷酸连接和检测测序系统(SOLID)3。该仪器将由麦克德莫特中心DNA测序核心实验室操作。最初，NGS基金将支持由NIH赞助的两项计划赠款资助的研究人员：由Michael Brown博士和Joseph Goldstein博士带头的项目项目(胆固醇代谢的分子基础)，该项目已进入资助的第32个年头；以及一项新的跨学科路线图赠款，即德克萨斯大学西南分校肥胖研究特别工作组，它汇集了来自16个不同部门的25个独立的PI。这项建议的结构是这样的：一个由13个拥有强大科学计划和进行具有生物学意义的观测记录的主要用户组成的小组将使用Solid 3系统来增强已经由NIH资助的实验。建议的研究包括使用基因表达谱来询问细胞对代谢扰动的反应，芯片测序来确定这一应用的PI发现的关键代谢转录因子的靶基因，以及DNA甲基化谱来评估饮食对表观遗传编程的影响。鉴于PI在这笔赠款上的科学生产力，我们乐观地认为，这项技术的应用将导致直接影响人类健康的重要新发现。虽然NGS设施的用户最初将被限制为13名调查人员，但这些科学家收集的经验将迅速传播到整个德克萨斯大学西南研究界。我们预计，在不久的将来，应用范围和用户范围都将扩大，Solid系统将成为麦克德莫特中心DNA测序核心的组成部分，十多年来，麦克德莫特中心DNA测序核心在德克萨斯大学西南分校提供了出色的大学服务。

DESCRIPTION (provided by applicant): Next-generation sequencing (NGS) provides a powerful new technology to rapidly and effectively interrogate DNA sequence in a simple, comprehensive and cost-effective manner. Probing patterns of whole genome expression, defining the full panoply of genes targeted by transcription factors, and assessing genome-wide methylation status has become feasible and affordable using NGS. The goal of this grant is to establish a high through-put NGS Facility to support ongoing research projects by a thematically-linked cadre of scientists who have a history of implementing new technologies to make high- impact, fundamental discoveries with direct applications to human health. The research focuses on two major causes of death and disability: disorders of cholesterol metabolism and dysregulated fuel homeostasis. Funds are requested to purchase a Sequencing by Oligonucleotide Ligation and Detection (SOLiD) 3 System from Applied Biosystems. The instrument will be operated by the McDermott Center DNA Sequencing Core Laboratory. Initially, the NGS Facility will support investigators funded by two NIH- sponsored programmatic grants: a Program Project (The Molecular Basis of Cholesterol Metabolism) spearheaded by Drs. Michael Brown and Joseph Goldstein that is in its 32nd year of funding, and a new interdisciplinary Roadmap grant, the Taskforce for Obesity Research at UT Southwestern, which brings together 25 independent PI's from 16 different departments. This proposal is structured so that a group of 13 major users with strong scientific programs and a track record of making biologically significant observations will use the SOLiD 3 System to enhance experiments already funded by the NIH. Proposed studies include using gene expression profiling to interrogate the cellular responses to metabolic perturbations, CHiP-sequencing to identify target genes of key metabolic transcription factors discovered by PI's of this application, and DNA methylation profiling to assess the impact of diet on epigenetic programming. Given the scientific productivity of the PI's on this grant, we are optimistic that application of this technology will result in important new discoveries that will directly impact human health. Although the users of the NGS Facility will initially be limited to 13 investigators, the experience gleaned by these scientists will be rapidly disseminated to the entire UT Southwestern research community. We anticipate that both the scope of applications and the spectrum of users will expand in the near future and that the SOLiD System will become an integral part of the McDermott Center DNA Sequencing Core that has provided excellent university-wide service at UT Southwestern for over a decade.