Author:
Shiokawa Mai,Miura Ryotaro,Okubo Aki,Hagita Yujiro,Yoshimura Itaru,Aoki Hiroshi
Abstract
AbstractBovine-derived cultured cells, including Madin-Darby bovine kidney cells, are used worldwide; however, lipofection tend to result in low transfection efficiency, which has impeded the progress of veterinary research. We performed experiments to confirm the lipofection efficiency of bovine-derived cultured cells, to identify cells that suitable for lipofection. Several bovine tissues (endometrium, testis, ear tissue and foetal muscle) were collected, and primary cultured cells were prepared. Lipofection assay showed that only bovine endometrium (BE)-derived cells could be transfected efficiently (50‒70%). BE cells can be divided into at least two types of cell populations (BE-1 and BE-2). The BE-1 cells, which were suitable for lipofection, were obtained by passages at short intervals and were negative for cytokeratin- and positive for vimentin-expression; the BE-2 cells did not have these characteristics and were not suitable for lipofection. Furthermore, the BE-1 cells and artificially immortalised cells of BE-1, iBE-1 cells, were utilised in a reporter assay requiring the introduction of multiple DNAs. Endometrial tissues can be collected from living cows, and BE-1 cells can be obtained easily by controlling passaging timing. The production of BE-1 cells and sharing the methods required to prepare them will contribute to the development of veterinary research.
Publisher
Springer Science and Business Media LLC
Reference30 articles.
1. Kim, T. K. & Eberwine, J. H. Mammalian cell transfection: The present and the future. Anal. Bioanal. Chem. 397, 3173–3178 (2010).
2. Vaheri, A. & Pagano, J. S. Infectious poliovirus RNA: A sensitive method of assay. Virology 27, 434–436 (1965).
3. McCutchan, J. H. & Pagano, J. S. Enchancement of the infectivity of simian virus 40 deoxyribonucleic acid with diethylaminoethyl-dextran. J. Natl. Cancer Inst. 41, 351–357 (1968).
4. Graham, F. L. & van der Alex, J. E. A new technique for the assay of infectivity of human adenovirus 5 DNA. Virology 52, 456–467 (1973).
5. Chen, C. & Okayama, H. High-efficiency transformation of mammalian cells by plasmid DNA. Mol. Cell Biol. 7, 2745–2752 (1987).