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Efficient transfection of primary zebrafish fibroblasts by nucleofection

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Abstract

Although various gene delivery techniques are available, their application in zebrafish cell cultures has not been extensively studied. Here, we report that nucleofection of zebrafish primary embryonic fibroblasts results in higher transfection efficiency in comparison to other non-viral gene delivery methods. The transfection was performed using green fluorescent protein (GFP) gene constructs of a different size. Greatest DNA uptake was obtained with 4.9-kb plasmid, resulting in 43% GFP positive cells. Nucleofection with 7.4-kb pH2B-GFP plasmid followed by geneticin (G418) selection was successfully used to establish a cell line expressing nuclear histone 2B-GFP fusion protein. Efficient transfection of zebrafish fibroblasts by nucleofection offers a non-viral technique of plasmid delivery and can be used to overexpress genes of interest in these cells.

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Acknowledgements

We acknowledge Mark T. Keating in whose laboratory the project was initiated and supported. We thank J. Chan, M. Lutchman, S. Arab and I. Splawski for critical reading of the manuscript, F. Engel for discussions and M. Schebasta for sharing plasmids. The financial support was provided by an NIH/NHLBI award no. P50 HLO74734–02 and SCOR in Pediatric Heart Development and Disease. A.J. acknowledges the Swiss National Science Foundation fellowship for advanced researchers.

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  1. Department of Cardiology, Children’s Hospital Boston, Enders 1261, 320 Longwood Avenue, Boston, MA, 02115, USA

    Rossen Badakov

  2. Department of Pediatrics, Harvard Medical School, Enders 1261, 320 Longwood Avenue, Boston, MA, 02115, USA

    Anna Jaźwińska

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  1. Rossen Badakov
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  2. Anna Jaźwińska
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Correspondence to Anna Jaźwińska.

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Badakov, R., Jaźwińska, A. Efficient transfection of primary zebrafish fibroblasts by nucleofection. Cytotechnology 51, 105–110 (2006). https://doi.org/10.1007/s10616-006-9018-3

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  • DOI: https://doi.org/10.1007/s10616-006-9018-3

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