(1118-C) Generating GFP-expressing CHO-K1 cell lines using the Tecan UNO Single Cell Dispenser

Abstract: Clonal isolation of single cells is a common method utilized in the field of Cell Biology to generate cell lines with specific genetic attributes for use across cellular, molecular, and biochemical applications. These genetic attributes can be used to explore cellular signaling pathways or simply to generate fluorescent tagged cells to be used for imaging or sorting applications. The Tecan Uno Single Cell Dispenser is a fast, efficient way to generate single cell clones with high fidelity and throughput for these types of applications. Here we use Uno for single cell isolation of CHO-K1 cells previously electro-transfected with a gene construct to express Green Fluorescent Protein (GFP) and selected with antibiotic to enrich for GFP expressing cells. We isolated a 384-well plate of single cells and found that 93% of wells contained a single cell. Following cell growth and expansion for 10 days, we observed 41.7% plating efficiency colony formation from GFP expressing single cells. We identified and selected three of the brightest GFP expressing clonal populations by fluorescent microscopy and expanded these to large format dishes. GFP stability was monitored during the expansion phase and evaluated again after cell stock freeze-thaw. The outcome of this work is CHO-K1 GFP-expressing monoclonal cell lines generated in 5 weeks. Broadly, these experiments demonstrate the utility of Uno for single cell cloning and cell line development.