Method for constructing kidney cell line of scophthalmus maximus

Abstract

The present invention discloses a method for constructing a kidney cell line of scophthalmus maximus, which comprises the following steps of: preparing a cell culture fluid first, taking a kidney tissue of the scophthalmus maximus as a material, using 0.5 percent trypsinase to digest the kidney tissue for 30 minutes at room temperature after the rinsing and shearing, adopting an 200-mesh nylon gauze to filter and collect cells, split charging the cells with the quantity of 5-10*10<5> counts / bottle into culture bottles with the growth area of 25 cm<2>, and placing the culture bottles at a temperature of 24 DEG C for culturing; replacing the cell culture fluid with half quantity each four days, and using 0.25 percent trypsinase digest the cells for passage when primary cells grow to be monolayered; and performing passage once each 8 to 10 days, wherein the content of serum in the cell culture fluid is reduced to 10 percent from 20 percent when the 8th generation is reached, and the cell line is successfully established at the moment. The kidney cell line of the scophthalmus maximus obtained by utilizing the method is fibroid and can be continuously transferred for more than 40 generations; the cell line can be directly applied to pathogeny characteristics research, vaccine development and functional gene research; and the construction method is suitable to construct kidney cell lines of other fishes.

Description

Technical field: [0001] The invention belongs to marine biological cell culture technology, and relates to a method for constructing a turbot kidney cell line using turbot kidney tissue. Background technique: [0002] The construction and cultivation of animal cell lines has long been a powerful tool for the study of virology, immunology, genetics, toxicology, and tumor therapy. Since the 1960s, more than 100 fish cell lines have been established and applied to the research of virology and immunology. [0003] Turbot (Scophthalmus maximus) is a rare fish species native to Europe. Since it was introduced into my country in 1992 and carried out industrialized farming, the annual output value of turbot has exceeded 4 billion yuan, and it has become a pillar industry of marine aquaculture in northern my country. However, with the rapid development of the turbot aquaculture industry, diseases caused by pathogens such as Vibrio and iridoviruses have become more and more frequent,...

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Problems solved by technology

[0004] So far, although there are turbot embryo cell lines (Yellow Sea Fisheries Research Institute of Chinese Academy of Fishery Sciences, Chen Songlin, Aquaculture249: 63-68), fin cell lines (Ocean University of China, Fan Tingjun, Journal of Ocean University of China 37 (5): 759-766) and a few cell lines reported in the literature, but there are still no reports of immune tissue such as spleen, kidney and other cell lines

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