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Establishment, characterization, and transfection potential of a new continuous fish cell line (CAM) derived from the muscle tissue of grass goldfish (Carassius auratus)

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Abstract

A new continuous fish cell line (CAM) has been successfully derived from the muscle tissues of grass goldfish, Carassius auratus. The primary cell cultures were initiated by incomplete trypsinization first and then explant culture in a Leibovitz-15 medium supplemented with 15% fetal bovine serum and 10% fish muscle extract. It was found that the CAM cells were very sensitive to trypsinization and needed to be sub-cultured at a low trypsin concentration of 0.0625% to be able to go through the crisis of spontaneous immortalization transformation, and afterward a total of five derivative cell strains were isolated from the original CAM cell line. This spontaneous immortalization transformation event was recorded successively at passages 44–47, beginning with a large-scale apoptosis and senescence and followed by mitosis arrest and re-activation, thus designated as cell strain CAM-44A, 44B, 45A, 44B, and 47A. Now both the CAM cell line and strains had been sub-cultured for more than 89 times and could be well cryopreserved in the growth medium containing 5% dimethylsulfoxide. Chromosome analysis and COI gene analysis had confirmed the grass goldfish origin of these CAM cells. Transfection potential analysis indicated that Lipofectamine LTX and Xfect were two suitable transfection reagents to be used in the gene delivery of CAM cells with a transfection efficiencies up to 11±6% and 8±3% in the CAM cell lines, respectively. Among the five cell strains, CAM-47A showed the highest transfection potential with a transfection efficiency up to 28 ± 5%. This work will provide a useful cell source for works on the cell-based artificial fish meat production and functional studies of fish myogenesis-related genes.

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Funding

This work was supported by the National Key R & D Program of China (Grant No. 2018YFD0901301), the Natural Science foundation of Shandong Province (ZR2020MC189), and the Fundamental Research Funds for Central Universities of China (Grant NO. 201822018).

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Authors and Affiliations

  1. Key Laboratory of Marine Genetics and Breeding, Ministry of Education, and College of Marine Life Sciences, Ocean University of China, Qingdao, 266003, China

    Na Li, Liwen Guo & Huarong Guo

  2. Institute of Evolution and Marine Biodiversity, Ocean University of China, Qingdao, 266003, China

    Huarong Guo

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  1. Na Li
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  2. Liwen Guo
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  3. Huarong Guo
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Contributions

Li N. carried out the work, processed the data and figures, and wrote the manuscript. Guo L. finished the analysis of transfection potential of CAM cells by Xfect. Guo H. designed the work and revised the manuscript.

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Correspondence to Huarong Guo.

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Li, N., Guo, L. & Guo, H. Establishment, characterization, and transfection potential of a new continuous fish cell line (CAM) derived from the muscle tissue of grass goldfish (Carassius auratus). In Vitro Cell.Dev.Biol.-Animal 57, 912–931 (2021). https://doi.org/10.1007/s11626-021-00622-1

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