CAREER: Revisiting the Central Dogma at the Genome Scale and in Single Cells

这个研究项目的最终目标是开发一个微设备平台，可以同时测量单个细胞的基因组、转录组、蛋白质组和表型参数，换句话说，在所有水平的中心规则（DNA到RNA到蛋白质到表型）。具体来说，PI将开发一种微流体装置，用于可靠地从同一单个细胞中分离细胞质和细胞核内容物，然后进行基因组dna和信使rna的全池扩增。扩增产物将被检索并使用最先进的基因组学技术进行分析，以将单细胞遗传变异与整个基因组中受影响基因的表达联系起来。单细胞、多重蛋白测定和细胞行为（表型）的定量分析可以集成在这个微设备中，以进一步将基因、转录本与蛋白质水平和细胞功能联系起来。PI将应用这个平台来研究干细胞分化，并可能为理解基因表达如何被调节来决定干细胞的命运和多样性提供新的见解。这种分析对于充分解决诸如胚胎发育、癌症和免疫细胞多样性等一系列基本生物学问题至关重要。该项目还作为从K12到研究生培训，从耶鲁大学到当地社区的研究和教育活动整合的支柱。将微技术融入生物学是未来健康科学研究人员和医疗从业者对专业知识的迫切需求。研究和教育的协同作用将导致虚拟微观生理系统实验室的发展，不仅向耶鲁大学的学生展示，也向公众展示，并增加少数民族和代表性不足群体的参与。由于该项目的跨学科性质，CBET部门的生物技术、生化和生物质工程项目的CAREER奖由分子和细胞生物学部门的系统和合成生物学项目共同资助。

1351443Fan, Rong The ultimate objective of this research project is to develop a microdevice platform that can enable the measurement of a single cell simultaneously for genomic, transcriptomic, proteomic and phenotypic parameters, in other words, at all the levels of the Central Dogma(DNA to RNA to protein to phenotype). Specifically, the PI will develop a microfluidic device for reliably separating cytoplasmic and nuclear contents from the same single cells followed by whole-pool amplification of genomic DNAs and messenger RNAs. The amplified products will be retrieved and analyzed using state-of-the-art genomics technology to link single-cell genetic variation to the expression of affected genes across the whole genome. Single-cell, multiplex protein assay and quantitative analysis of cellular behaviors (phenotype) can be integrated within this microdevice to further correlate genes, transcripts to the levels of proteins and cellular functions. The PI will apply this platform to investigating stem cell differentiation and potentially provide new insights needed for understanding how gene expression is regulated to determine stem cell fate and diversity. Such analysis is critical to fully addresse a range of fundamental biological questions such as embryonic development, cancer and immune cell diversity. This project also acts as a backbone for the integration of research and educational activities spanning from K12 through graduate training, from Yale University through the local community. Integrating microtechnology into biology represents an urgent requirement of expertise for the future health science researchers and medical practitioners. The synergy of research and education will result in the development of a virtual micro-physiologic system lab for demonstration to not only Yale College students but also to the general public, and increase the participation of minorities and under-represented groups.Due to the interdisciplinary nature of the project, this CAREER award by the Biotechnology, Biochemical, and Biomass Engineering Program of the CBET Division is co-funded by the Systems and Synthetic Biology Program of the Division of Molecular and Cellular Biology.