High-throughput micro-RNA profiling of single cells and its application in leukemia

项目摘要 单细胞RNA测序（scRNA-seq）有望彻底改变生物学研究 通过允许测量单个细胞内的细胞过程， 精确度和广度。micro-RNA（miRNA）是一类小的调控RNA， 参与一系列生物过程，如有机体发育、细胞 分化和主要信号传导途径。miRNA特征与疾病密切相关 表型，可以作为可靠的生物标志物。 该提案将开发一种微流体平台，可以允许分离和测序。 以高度平行和无偏的方式从单个单细胞中提取miRNA。在K99阶段，我 将按顺序开发装置的每个部分，并优化反应和过程 将使用对照细胞系对该装置进行测试和验证，以确定与其相关的生物学特性。 检测的灵敏度，并建立单细胞数据的质量。最后，我将使用平台 分析白血病细胞系和从患者获得的细胞系中的miRNA表达， 以鉴定具有诊断或预后价值的表达模式。 该项目的R 00阶段将侧重于将该平台应用于研究疾病 白血病的发病机制这项研究的结果将揭示疾病的新途径 进展和耐药性，可用于开发新的治疗方式。我 还将开发该平台作为检测白血病的超灵敏诊断工具， 患者 我还提出了一个广泛的培训计划，这将有助于我实现我的职业目标和支持 我向独立研究职位的过渡科赫研究所的研究环境是 无与伦比的，并提供了独特的机会，互动和合作，与著名的生物学家 和工程师。我召集了一个特殊的团队：丹安德森教授和罗伯特教授 兰格是生物工程和高通量技术方面的专家，他将指导我。 我与RNA生物学家菲利普夏普教授密切合作，他将帮助我解决分子生物学问题， 项目的组成部分。Amir Faithi博士将提供临床样本和指导， 该项目的医疗影响。我的培训还包括指导学生，展示我的 在单细胞会议和科学推广中工作。

Project Summary Single-cell RNA sequencing (scRNA-seq) holds the promise to revolutionize the study of biology by allowing measurement of cellular processes within individual cells with unprecedented precision and breadth. Micro-RNAs (miRNA) are a class of small regulatory RNAs that are involved in a range of biological processes such as organismal development, cellular differentiation, and major signaling pathways. miRNA signatures correlate strongly with disease phenotype and can serve as reliable biomarkers. This proposal will develop a microfluidic platform that can allow isolation and sequencing of miRNA from individual single-cells in a highly-parallel and unbiased manner. In the K99 phase, I will sequentially develop each segment of the device and optimize the reactions and processes associated with it. The device will be tested and validated using control cell lines to determine the sensitivity of detection, and establish the quality of single cell data. Finally, I will use the platform to profile miRNA expressions in leukemia cell lines and those obtained from patients with the goal to identify expression patterns that will have diagnostic or prognostic value. The R00 phase of the project will focus on applying this platform in studying disease pathogeneses in leukemia. Findings from the study will shine light on new pathways for disease progression and drug resistance, which can be used for developing new therapeutic modalities. I will also develop the platform as an ultra-sensitive diagnostic tool for detecting leukemia in patients. I also propose an extensive training program that will help me realize my career goals and support my transition to an independent research position. The research environment at Koch Institute is unparalleled and offers the unique opportunity to interact and collaborate with eminent biologists and engineers. I have assembled an exceptional team: Prof. Dan Anderson and Prof. Robert Langer, experts in bioengineering and high-throughput techniques, will mentor me. I will work very closely with Prof. Phillip Sharp, RNA Biologist, who will help me address the molecular biology component of the project. Dr. Amir Faithi will provide clinical samples and direction to increase the medical impact of the project. My training will also involve mentoring students, presenting my work in single-cell meetings, and science outreach.