A Split-Luciferase-Based Cell Fusion Assay for Evaluating the Myogenesis-Promoting Effects of Bioactive Molecules
- PMID: 34050464
- DOI: 10.1007/978-1-0716-1258-3_8
A Split-Luciferase-Based Cell Fusion Assay for Evaluating the Myogenesis-Promoting Effects of Bioactive Molecules
Abstract
A split-luciferase-based cell fusion assay enables high-throughput screening of myogenesis-promoting chemicals in chemical libraries. The assay consists of two C2C12 myoblast-derived cell lines (N- and C-cells), each of which stably expresses either an N- or C-terminal split-firefly luciferase (FLuc) fragment fused to a naturally split DnaE intein (N- and C-probes, respectively). The fusion of N- and C-cells during myogenesis induces bioluminescence (BL) in the cytosol due to a stable reconstitution of the split-FLuc. Thus, the myogenesis-promoting effects of a chemical compound can be determined through the enhanced BL intensity. Here, we describe the preparation of N- and C-cells and determination of the myogenesis-promoting effects of imatinib using a 96-well microplate-based assay.
Keywords: 96-Well microplate-based assay; C2C12 myoblast; Cell fusion; DnaE intein; Firefly luciferase; Imatinib; Myogenesis; Protein splicing; Split luciferase reconstitution.
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