Genetic analysis of single cells

生物学研究中一个基本且未解决的挑战是需要了解和测量单个细胞的基因组和转录状态。例如，地球上几乎所有的微生物都没有被培养过，而且由于必须与其他生物建立共生关系，它们很可能难以在实验室环境中培养。因此，这些生命形式不适合目前的基因组研究技术，因为基因组研究需要大量的纯细胞群。另一方面，即使是基因相同的细胞，如人体内的细胞，由于分化状态、随机变异和疾病，也表现出广泛的异质性。可扩展的单细胞分析技术是迫切需要的，它将对包括微生物多样性、癌症生物学和人类发展在内的各个领域产生深远的影响。尽管目前在仪器和化学方面取得了进展，可用于全球测序和敏感的靶向转录测量，但这些都受到操纵单细胞的困难和传统大体积（μ L）样品处理产生的低效率的限制。在这里，我们将把这些成熟技术的力量与制造高度集成的“片上实验室”设备的新工程方法结合起来，开发能够以更高的灵敏度和速度处理数百到数千个单细胞的微系统。具体来说，我们将开发两种技术：1)从环境样本中提取的未培养微生物的高通量分离和基因组扩增，以及ii)哺乳动物细胞中miRNA表达的可扩展分析。

A fundamental and unmet challenge in biological research is the need to understand and measure the genome and transcriptional state of individual cells. For instance, nearly all microorganisms on the planet have not been cultivated and may very well be refractory to culture in laboratory settings due to requisite symbiotic relationships with other organisms. Thus, these life forms are not amenable to current technologies for genomic study, which require large numbers and pure populations of cells. On the other hand, even genetically identical cells, such as those in the human body, exhibit widespread heterogeneity owing to differentiation state, stochastic variations, and disease. Scalable single cell analysis technologies are urgently needed and will have far reaching impact touching diverse fields including microbial diversity, cancer biology, and human development. Although advances in instrumentation and chemistries for global sequencing and sensitive targeted measurements of transcription are now available, these are limited by difficulties in manipulating single cells and inefficiencies arising from conventional large-volume (µL) sample processing. Here we will couple the power of these established technologies with new engineering methods for the fabrication of highly integrated "lab-on-a-chip" devices to develop microsystems capable of processing hundreds to thousands of single cells with improved sensitivity and speed. Specifically, we will develop two technologies for i) the high-throughput isolation and genome amplification of uncultivated microbes derived from environmental samples, and ii) the scalable analysis of miRNA expression in mammalian cells.