An optimized FusX assembly-based technique to introduce mitochondrial TC-to-TT variations in human cell lines

• DOI: 10.1016/j.xpro.2022.101288
• Publication Date: 2022-04-14T03:15:40Z
• Authors: Bibekananda Kar; Ankit Sabharwal; Santiago Restrepo...; Brandon W. Simone; Karl J. Clark; Stephen C. Ekker
• Keywords: 0303 health sciences; Science (General); Cell Biology; DNA, Mitochondrial; Cell Line; Mitochondria; Q1-390; 03 medical and health sciences; CRISPR; Genetics; Protocol; Sequencing; Humans; Molecular Biology

The FusX TALE Based Editor (FusXTBE) is a programmable base editing platform that can introduce specific TC-to-TT variations in the mitochondrial DNA (mtDNA). Here, we provide a protocol describing the synthesis and testing of the FusXTBE plasmids in cultured human cell lines. This tool is designed to be easily modified to work in diverse applications where editing of mitochondrial DNA is desired. For complete details on the use and execution of this protocol, please refer to Sabharwal et al. (2021) and Ma et al. (2016).