Microfluidics-Based Single-Cell Chemical Analysis of Cyanobacteria

单细胞分析是一项新兴技术，有望为生物学提供新的视角。因为单细胞分析将细胞作为个体来观察，它消除了总体平均，并解决了样本群体中存在的异质性。这种异质性可以提供有关重要细胞过程的有用信息，特别是如果相当多的细胞表现出明显偏离平均细胞行为的情况。本研究项目将在微流控平台上开发单细胞分析技术，以研究单细胞蓝藻聚球藻的藻胆体(PBS)降解过程。通过单分子检测，可以直接从单细胞裂解液中计算出非常低的荧光分子拷贝数，漂白的蓝藻细胞就是这种情况。这些工具将被用来在单细胞水平上了解PBS降解的生化机制。广泛的影响该项目将对许多不同的研究领域产生重大影响。首先，从这项研究中获得的生物学洞察力将阐明看待生态多样性、微生物学以及它们之间相互关系的新方法。第二，单细胞技术将朝着细胞培养、细胞分离和操作以及细胞内内容物(蛋白质和核酸)分析的一体化方向发展。使用这种综合方法，应该可以从单个细胞获得准确的生化信息以及简单的表型变化。这一策略可以推广到其他类型的细胞，并应该为研究生物异质性提供一个新的范式。第三，这个项目本质上是跨学科的，涉及化学分析和生物结果。这种结合应该会导致许多领域之间的协同对话，包括生态学、种群生物学和生物物理化学，从而为未来的研究带来更有成效的方向和动机。第四，将在本科生和研究生一级进行强有力的教学和培训，以及每年举办多学科讲习班，向参与者介绍单细胞技术在生物学中的潜在用途。

Single-cell analysis is an emerging technology that promises to provide new perspectives on biology. Because single-cell analysis observes cells as individuals, it eliminates ensemble averaging and resolves the heterogeneity present within a sample population. This heterogeneity can provide useful information with regard to important cellular processes, especially if a significant number of cells display a marked deviation from that of average-cell behavior. This research project will be to develop single-cell analysis technology on a microfluidic platform to study the phycobilisome (PBS) degradation process of a unicellular cyanobacterium Synechococcus. With the single-molecule detection, very low copy numbers of fluorescent molecules can be directly counted from a single-cell lysate, which is the case for bleached cyanobacteria cells. These tools will be used to understand the biochemical mechanism of PBS degradation at the single-cell level.Broader ImpactsThis project will have a significant impact on many different areas of research. First, the biological insight gained from this research will illuminate a new way of viewing ecological diversity, microbiology, and the interrelationship between them. Second, single-cell technology will be advanced toward the integration of cell culture, isolation and manipulation of cells, and the analysis of intracellular contents (both proteins and nucleic acids). With this integrative approach, it should be possible to obtain accurate biochemical information from individual cells as well as simple phenotypic changes. This strategy can be extended to other types of cells and should provide a new paradigm for studying biological heterogeneity. Third, this project is interdisciplinary in essence, involving chemical analysis with biological outcomes. This combination should induce a synergistic conversation between many fields, including ecology, population biology, and biophysical chemistry, resulting in more productive directions and motivations for future research. Fourth, there will be a strong component of teaching and training both at the undergraduate and graduate level as well as annual multidisciplinary workshops to introduce participants to the potential uses of single-cell technology in biology.