Yuan Jinzhou, Zhou Jessie, Raizen David M, Bau Haim H
Department of Mechanical Engineering and Applied Mechanics, University of Pennsylvania, Philadelphia, PA 19104, USA.
Lab Chip. 2015 Jul 7;15(13):2790-8. doi: 10.1039/c5lc00305a. Epub 2015 May 26.
Animal motility varies with genotype, disease, aging, and environmental conditions. In many studies, it is desirable to carry out high throughput motility-based sorting to isolate rare animals for, among other things, forward genetic screens to identify genetic pathways that regulate phenotypes of interest. Many commonly used screening processes are labor-intensive, lack sensitivity, and require extensive investigator training. Here, we describe a sensitive, high throughput, automated, motility-based method for sorting nematodes. Our method is implemented in a simple microfluidic device capable of sorting thousands of animals per hour per module, and is amenable to parallelism. The device successfully enriches for known C. elegans motility mutants. Furthermore, using this device, we isolate low-abundance mutants capable of suppressing the somnogenic effects of the flp-13 gene, which regulates C. elegans sleep. By performing genetic complementation tests, we demonstrate that our motility-based sorting device efficiently isolates mutants for the same gene identified by tedious visual inspection of behavior on an agar surface. Therefore, our motility-based sorter is capable of performing high throughput gene discovery approaches to investigate fundamental biological processes.
动物的运动能力因基因型、疾病、衰老和环境条件而异。在许多研究中，进行基于运动能力的高通量分选以分离稀有动物是很有必要的，这尤其用于正向遗传筛选，以识别调控感兴趣表型的遗传途径。许多常用的筛选过程劳动强度大、缺乏灵敏度，并且需要研究人员进行大量培训。在此，我们描述了一种用于分选线虫的灵敏、高通量、自动化的基于运动能力的方法。我们的方法在一个简单的微流控装置中实现，该装置每个模块每小时能够分选数千只动物，并且易于并行操作。该装置成功地富集了已知的秀丽隐杆线虫运动突变体。此外，利用这个装置，我们分离出了能够抑制flp - 13基因催眠作用的低丰度突变体，flp - 13基因调控秀丽隐杆线虫的睡眠。通过进行遗传互补试验，我们证明基于运动能力的分选装置能够有效地分离出通过在琼脂表面对行为进行繁琐的目视检查所鉴定的同一基因的突变体。因此，我们基于运动能力的分选仪能够进行高通量基因发现方法来研究基本的生物学过程。
