Activity of Individual Aquatic Bacteria by Flow Cytometry; Instrumentation and Protocols

该奖项将支持流式细胞仪仪器和相关协议的开发，这些仪器和协议可以分析水生细菌，重点是它们的生长和活力状态。流式细胞术已经成为研究血细胞和各种细胞以及可以在培养物中生长的生物体的既定研究工具。 然而，其用于分析天然细菌种群，例如在水生系统中发现的那些细菌，由于这种细菌典型的小细胞尺寸而变得复杂。很少有商业仪器专门用于探测这些小目标。该项目的主要工作是设计对商业流式细胞仪的额外修改，以提高其确定水生细菌的生长和存活状态的能力。将对输出信号进行组织处理，以反映通过DNA含量判断的混合种群的生长速率。将在系统中检测另外两种活性探针，一种用于RNA含量，另一种用于膜电位，这两种探针都是细菌生长和活力的重要指标。该设备对于研究高纬度地区的微生物产生温室气体特别重要，因为冰盖和细菌活动对温度高度敏感。

This award will support the development of flow cytometry instrumentation and associated protocols that can analyze aquatic bacteria with emphasis on their growth and viability status. Flow cytometry has become an established research tool for study of blood cells and of a variety of cells and organisms that can be grown in culture. However, its use to analyze natural populations of bacteria such as those found in aquatic systems is complicated by the small cell size typical of such bacteria. Few commercial instruments are dedicated to the detection of these small targets. The main effort of this project is to design additional modifications to a commercial flow cytometer to enhance its ability to determine the growth and viability status of aquatic bacteria. Output signals will be organized for manipulation to reflect the rate of growth in mixed populations as judged by DNA content. Two additional probes for activity will be tested in the system, one for RNA content and the other for membrane potential, both important indicators of bacterial growth and viability. The equipment will be particularly important in study of the microbial generation of greenhouse gasses at high latitudes where both ice cover and bacterial activity is highly temperature sensitive.