黑点青鳉鱼(Oryziasmelastigma)鳍上皮细胞系的建立及其特性分析
首发时间:2019-03-07
摘要:黑点青鳉鱼(Oryzias melastigma)是一种理想的海洋模式生物,且其全基因组测序已完成。本研究通过用胰蛋白酶处理黑点青鱂鱼的鳍上皮组织进行原代细胞培养。获得的原代细胞呈铺路石状,与上皮细胞的特征相符。常温条件下能在含15%胎牛血清、1%罗非鱼血清、10 ng/ml 的碱性成纤维生长因子和100 μM β -巯基乙醇的DMEM/F12培养基中生长繁殖。通过继代,这些细胞能在仅含15%胎牛血清的DMEM/F12培养基中传代培养,目前已连续培养了50多代,长势良好。染色体核型分析表明该细胞的染色体数目为48条;现已成功地将携带绿色荧光蛋白编码基因的重组真核表达载体pcDNA/GFP质粒转染入该黑点青鳉鱼的细胞中,荧光显微镜观察显示绿色荧光蛋白基因已在该细胞系中成功表达,证明该细胞系可用于外源基因的真核表达以及外源基因的功能研究。同时成功地用该细胞系感染鱼的神经坏死病毒,证明了黑点青鳉鱼鳍上皮细胞系可作为病毒的宿主细胞,从而进行鱼类疾病的研究。因此,本项研究将为海水硬骨鱼类的细胞分子生物学、免疫学、毒理学等方面的研究奠定基础。
关键词: 生物学 青鳉鱼 鳍细胞系 原代培养 染色体核型分析
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Establishment and characterization of a cell line from marine medaka Oryziasmelastigma fin epithelium
Abstract: Marine medaka Oryzias melastigma is an ideal marine model organism with the completed genome sequencing. To provide a cell model for studies on the molecular biology, immunology, and ecotoxicology of marine teleosts, a cell line was established from the tissue of O. melastigma fins. The cells primarily looked like paving stone morphologically and multiplied well in DMEM/F12 medium supplemented with 15% fetal bovine serum, 1% fish serum and the growth factor at room temperature. Subsequently, they were subcultured in DMEM/F12 medium plus 15% fetal bovine serum only, and grew well. So far, they had been maintained for more than 50 subcultivations. Chromosome karyotyping revealed that the modal chromosome number in the cells was 48. In the cells transfected with plasmid pcDNA/GFP, green fluorescence was observed, which suggested that the cell line could be used to express foreign gene for the study on gene functions. Moreover, viral nervous necrosis virus (VNNV) gene was detected in the cells exposed to the virus, which implied that O. melastigma fin epithelial cell line might be used as the host cells for the study of viral nervous necrosis.
Keywords: biology Oryzias melastigma fin cell line primary culture chromosome karyotyping
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黑点青鳉鱼(Oryziasmelastigma)鳍上皮细胞系的建立及其特性分析
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