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In vitro development of porcine transgenic nuclear-transferred embryos derived from newborn Guangxi Bama mini-pig kidney fibroblasts

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Abstract

Porcine transgenic cloning has potential applications for improving production traits and for biomedical research purposes. To produce a transgenic clone, kidney fibroblasts from a newborn Guangxi Bama mini-pig were isolated, cultured, and then transfected with red and green fluorescent protein genes using lipofectamine for nuclear transfer. The results of the present study show that the kidney fibroblasts exhibited excellent proliferative capacity and clone-like morphology, and were adequate for generation of somatic cell nuclear transfer (SCNT)-derived embryos, which was confirmed by their cleavage activity and blastocyst formation rate of 70.3% and 7.9%, respectively. Cells transfected with red fluorescent protein genes could be passed more than 35 times. Transgenic embryos cloned with fluorescent or blind enucleation methods were not significantly different with respect to cleavage rates (92.5% vs. 86.8%, p > 0.05) and blastocyst-morula rates (26.9% vs. 34.0%, p > 0.05), but were significantly different with respect to blastocyst rates (3.0% vs. 13.2%, p < 0.05). Cleavage (75.3%, 78.5% vs. 78.0%, p > 0.05), blastocyst (14.1%, 16.1% vs. 23.1%, p > 0.05) and morula/blastocyst rates (43.5%, 47.0% vs. 57.6%, p > 0.05) were not significantly different between the groups of transgenic cloned embryos, cloned embryos, and parthenogenetic embryos. This indicates that long-time screening by G418 caused no significant damage to kidney fibroblasts. Thus, kidney fibroblasts represent a promising new source for transgenic SCNT, and this work lays the foundation for the production of genetically transformed cloned Guangxi Bama mini-pigs.

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Acknowledgments

We thank Dr. Bernard A. Goodman for his critical suggestions and revisions of the manuscript. This research was funded by grants from the National Natural Science Foundation of China (No. 31260553).

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The authors declare no conflict of interest.

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  1. State Key Laboratory for Conservation and Utilization of Subtropical Agro-Bioresources, and College of Animal Science and Technology, Guangxi University, 100 Daxuedong Road, Nanning, 530004, China

    Hongbo Liu, Peiru Lv, Xiangxing Zhu, Xianwei Wang, Xiaogan Yang, Erwei Zuo, Yangqing Lu, Shengsheng Lu & Kehuan Lu

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  1. Hongbo Liu
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  2. Peiru Lv
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Correspondence to Shengsheng Lu or Kehuan Lu.

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Liu, H., Lv, P., Zhu, X. et al. In vitro development of porcine transgenic nuclear-transferred embryos derived from newborn Guangxi Bama mini-pig kidney fibroblasts. In Vitro Cell.Dev.Biol.-Animal 50, 811–821 (2014). https://doi.org/10.1007/s11626-014-9776-8

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  • DOI: https://doi.org/10.1007/s11626-014-9776-8

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