Abstract

Bovine-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.

Highlights

  • Bovine-derived cultured cells, including Madin-Darby bovine kidney cells, are used worldwide; lipofection tend to result in low transfection efficiency, which has impeded the progress of veterinary research

  • Bovine primary cultures were derived from the bovine endometrium (BE), bovine testis (BT), ear tissue [as bovine fibroblasts (BF)] and bovine foetal muscle (BFM)

  • Cultured 293 T cells, which are known to be suitable for efficient gene ­introduction[15], showed high lipofection efficiency (64.8% ± 8.5%); the efficiency was low in BT (10.0% ± 6.1%), BF (6.1% ± 3.0%), BFM (2.3% ± 1.7%) and Madin-Darby bovine kidney (MDBK) cells (5.3% ± 1.0%)

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Summary

Introduction

Bovine-derived cultured cells, including Madin-Darby bovine kidney cells, are used worldwide; lipofection tend to result in low transfection efficiency, which has impeded the progress of veterinary research. The lipofection efficiency of cultured cells is often described in the product information provided by the manufacturers of lipofection reagents, allowing the transfection efficiency of targeted cells to be known prior to performing a lipofection This information is often limited to cultured cells that are used frequently, especially human- and rodent-derived cultured cells. Transfection assays were performed using cultured cells derived from bovine fibroblasts; no consensus has been reached regarding the lipofection efficiency in these ­cells[11,12,13] This low transfection efficiency in bovine-derived cultured cells makes experimentation difficult, especially those that require the introduction of multiple exogenous DNA sequences, such as reporter assays and genome editing strategies, hindering the progress of veterinary research. Producing bovine-derived cultured cells that solve the difficulties of lipofection and provide a method for their production This discovery will greatly contribute to the development of veterinary research

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