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，以简单、全面和具有成本效益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以及评估全基因组甲基化状态已变得可行且负担得起。这笔赠款的目标是建立一个高通量的NGS设施，以支持正在进行的研究项目，这些项目由具有实施新技术历史的科学家组成，这些科学家具有直接应用于人类健康的高影响力，基础性发现。该研究集中在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。 申请资金用于从应用生物系统公司购买寡核苷酸连接和检测测序（SOLiD）3系统。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设施将支持由两个NIH赞助的计划赠款资助的研究人员：一个由Michael Brown和Joseph Goldstein博士牵头的计划项目（胆固醇代谢的分子基础），这是其第32年的资助，以及一个新的跨学科路线图拨款，UT西南部肥胖研究工作组，汇集了来自16个不同部门的25个独立PI。 该提案的结构是这样的，一组13个拥有强大科学计划和进行生物学重要观察的记录的主要用户将使用SOLiD 3系统来加强已经由NIH资助的实验。提出的研究包括使用基因表达谱来询问对代谢扰动的细胞响应，CHiP测序来鉴定由本申请的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。考虑到PI在这项资助下的科学生产力，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设施的用户最初将限于13名研究人员，但这些科学家收集的经验将迅速传播到整个UT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测序核心的一个组成部分，该核心在UT西南部提供了良好的全校服务十多年来。

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.