Introduction of a plasmid and a protein into bovine and swine cells by water-in-oil droplet electroporation

摘要

Instrument cost is a major problem for the transduction of DNA fragments and proteins into cells. Water-in-oil droplet electroporation (droplet-EP) was recently invented as a low-cost andeffective method for the transfection of plasmids into cultured human cells. We here applied droplet-EP to livestock animal cells. Although it is difficult to transfect plasmids into bovinefibroblasts using conventional lipofection methods, droplet-EP enabled us to introduce an enhanced green fluorescent protein (EGFP)-expressing plasmid into bovine earlobe fibroblasts. Theoptimal transfection condition was 3.0 kV, which allowed 19.1% of the cells to be transfected. For swine earlobe fibroblasts, the maximum transfection efficacy was 14.0% at 4.0 kV. Aftertransfection with droplet-EP, 69.1% of bovine and 76.5% of swine cells were viable. Furthermore, droplet-EP successfully transduced recombinant EGFP intofrozen-thawed bovine sperm at 1.5 kV. Flow cytometry analysis revealed that 71.5% of spermatozoa exhibited green fluorescence after transfection. Overall, droplet-EP is suitable for thetransfection of plasmids and proteins into cultured livestock animal cells.