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)有望给生物学研究带来革命性的变化。 通过史无前例地测量单个细胞内的细胞过程 精确度和广度。Micro-RNAs(MiRNA)是一类小的调节RNA，它是 参与一系列生物过程，如生物发育、细胞 分化，以及主要的信号通路。MiRNA特征与疾病密切相关 表型，可作为可靠的生物标志物。 这项提议将开发一种微流控平台，可以分离和测序 以高度平行和无偏见的方式从单个单细胞中提取miRNA。在K99阶段，我 将按顺序开发设备的每个部分，并优化反应和过程 与之相关的。该装置将使用控制细胞系进行测试和验证，以确定 检测灵敏度，并建立单细胞数据的质量。最后，我将使用该平台 分析白血病细胞系和从患者那里获得的miRNA的表达 以确定将具有诊断或预后价值的表达模式。 项目的R00阶段将专注于将该平台应用于疾病研究 白血病的发病机制。这项研究的发现将为疾病的新途径带来曙光 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 病人。 我还提出了一个广泛的培训计划，帮助我实现我的职业目标和支持 我过渡到一个独立的研究职位。科赫研究所的研究环境是 无与伦比，并提供了与著名生物学家互动和合作的独特机会 和工程师。我组建了一个杰出的团队：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，他们将指导我。我会非常努力地工作 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 项目的组件。Amir Faithi博士将提供临床样本和增加方向 该项目的医疗影响。我的培训还将包括指导学生，介绍我的 在单细胞会议和科学外展中工作。

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.