Constructing method and application of koi brain cell line

Abstract

The invention discloses a constructing method and application of a koi brain cell line. The constructing method includes: placing koi brain tissue in digestive liquid A containing collagenase and trypsin for digestion; collecting cells; adding into digestive liquid b containing EDTA-4Na for digestion; performing primary culture and secondary culture after the cells which are digested are collected to obtain the koi brain cell line. Aiming at features of koi brain tissue cells, a digestion method combining collagenase with trypsin is adopted to separate tissue cells for primary culture, the koi brain cell line is constructed successfully and continuously transferred to more than 100 generations already, and a lot of koi brain source cells can be provided; the cells can maintain good growing state, so that the cells can be freeze-stored. The koi brain cell line is sensitive to koi herpes viruses with cytopathy occurring after being exposed to the viruses, and proliferated virus particles can be found through electron microscope observation. And the koi brain cell line can be directly applied to pathogenic characteristic study and vaccine development.

Description

technical field [0001] The invention relates to a method for constructing a koi brain cell line and its application in isolating viruses, belonging to freshwater biological cell culture application technology. Background technique [0002] Koi ( Cyprinus carpio ) is also known as "Fei carp" and "Japanese koi", which is one of the large ornamental fish and is a variety of carp. , Showa tricolor and other 13 strains. Because of their good size and bright colors, the scale of breeding of koi is increasing. Along with breeding environment more and more bad, the various infectious diseases of koi especially koi herpes virus disease begin to spread widely. Koi herpes virus disease first broke out in Israel in 1998, but it was confirmed in 1999 that its real pathogen was Koi herpes virus (KHV). Since then, outbreaks have also occurred in the United Kingdom, continental Europe, the United States, and Japan. In recent years, the widespread spread of koi herpes virus disease has ...

AI Technical Summary

Problems solved by technology

Digestion method is currently commonly used trypsin digestion, but trypsin will cause damage to some epithelial cells, and will not be effective on fibers and connective tissues

So far, the literature reports on the preparation of koi cell lines by enzymatic digestion include: Dong Chuanfu, Xue Shuqun and Xiao Yi all established koi fin ray cell lines by using the tissue block separation method (VirusResearch, 2011, 161: 140-149; Fisheries Science Journal, 2012, 25 (4): 16-23; Chinese Journal of Cell Biology 2012, 34 (8): 767-774), Wang Yingying established a Koi snout cell line by digestion method (Aquaculture, 2015, 435 : 310–317), but there are no reports of koi brain-derived cell lines

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