Enhanced transfection of cell lines from Atlantic salmon through nucoleofection and a
Enhanced transfection of cell lines from Atlantic salmon through nucoleofection and antibiotic selection
Abstract Background Cell lines from Atlantic salmon kidney have made it possible to culture and study infectious salmon anemia virus (ISAV), an aquatic orthomyxovirus affecting farmed Atlantic salmon. However, transfection of these cells using calcium phosphate precipitation or lipid-based reagents shows very low transfection efficiency. The Amaxa Nucleofector technology™ is an electroporation technique that has been shown to be efficient for gene transfer into primary cells and hard to transfect cell lines.
Findings Here we demonstrate, enhanced transfection of the head kidney cell line, TO, from Atlantic salmon using nucleofection and subsequent flow cytometry. Depending on the plasmid promoter, TO cells could be transfected transiently with an efficiency ranging from 11.6% to 90.8% with good viability, using Amaxa's cell line nucleofector solution T and program T-20. A kill curve was performed to investigate the most potent antibiotic for selection of transformed cells, and we found that blasticidin and puromycin were the most efficient for selection of TO cells.
Conclusions The results show that nucleofection is an efficient way of gene transfer into Atlantic salmon cells and that stably transfected cells can be selected with blasticidin or puromycin.
- Content Type Journal Article
- Pages 1-7
- DOI 10.1186/1756-0500-4-136
- Berit L Schiøtz, Department of Pharmaceutical Biosciences, School of Pharmacy, University of Oslo, Oslo, Norway
- Esther G Rosado, Department of Microbiology, Faculty of Science, University of Malaga, Malaga, Spain
- Espen S Baekkevold, Institute of Pathology, Rikshospitalet-Radiumhospitalet Medical Center, University of Oslo, Oslo, Norway
- Morten Lukacs, Department of Basic Science and Aquatic Medicine, Norwegian School of Veterinary Science, Oslo, Norway
- Siri Mjaaland, Department of Food Safety and Infection Biology, Norwegian School of Veterinary Science, Oslo, Norway
- Hilde Sindre, National Veterinary Institute, Oslo, Norway
- Unni Grimholt, Centre for Ecology and Evolutionary Synthesis, Department of Biology, University of Oslo, Oslo, Norway
- Tor Gjøen, Department of Pharmaceutical Biosciences, School of Pharmacy, University of Oslo, Oslo, Norway