Funano Shun-Ichi, Tone Daisuke, Ukai Hideki, Ueda Hiroki R, Tanaka Yo
Laboratory for Integrated Biodevice, Center for Biosystems Dynamics Research, RIKEN, 1-3 Yamadaoka, Suita, Osaka, 565-0871, Japan.
Laboratory for Synthetic Biology, Center for Biosystems Dynamics Research, RIKEN, 1-3 Yamadaoka, Suita, Osaka, 565-0871, Japan.
BMC Res Notes. 2020 Oct 5;13(1):453. doi: 10.1186/s13104-020-05294-w.
Production of genetically modified mice including Knock-out (KO) or Knock-in (KI) mice is necessary for organism-level phenotype analysis. Embryonic stem cell (ESC)-based technologies can produce many genetically modified mice with less time without crossing. However, a complicated manual operation is required to increase the number of ESC colonies. Here, the objective of this study was to design and demonstrate a new device to easily find colonies and carry them to microwells.
We developed a polydimethylsiloxane-based device for easy manipulation and isolation of ESC colonies. By introducing ESC colonies into the groove placed near culturing microwells, users can easily find, pick up and carry ESC colonies to microwells. By hydrophilic treatment using bovine serum albumin, 2-μL droplets including colonies reached the microwell bottom. Operation time using this device was shortened for both beginners (2.3-fold) and experts (1.5-fold) compared to the conventional colony picking operation. Isolated ESC colonies were confirmed to have maintained pluripotency. This device is expected to promote research by shortening the isolation procedure for ESC colonies or other large cells (e.g. eggs or embryos) and shortening training time for beginners as a simple sorter.
为了进行生物体水平的表型分析，生产基因编辑小鼠，包括基因敲除（KO）或基因敲入（KI）小鼠是必要的。基于胚胎干细胞（ESC）的技术可以在不进行杂交的情况下，用更少的时间生产出许多基因编辑小鼠。然而，需要复杂的手动操作来增加胚胎干细胞集落的数量。本研究的目的是设计并展示一种新装置，以便轻松找到集落并将其转移到微孔中。
我们开发了一种基于聚二甲基硅氧烷的装置，用于轻松操作和分离胚胎干细胞集落。通过将胚胎干细胞集落引入培养微孔附近的凹槽中，用户可以轻松找到、拾取并将胚胎干细胞集落转移到微孔中。通过使用牛血清白蛋白进行亲水处理，包含集落的2微升液滴到达了微孔底部。与传统的集落挑选操作相比，使用该装置的操作时间对于初学者缩短了2.3倍，对于专家缩短了1.5倍。经确认，分离出的胚胎干细胞集落保持了多能性。作为一种简单的分选器，该装置有望通过缩短胚胎干细胞集落或其他大细胞（如卵子或胚胎）的分离程序以及缩短初学者的培训时间来促进研究。
