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Carp spinal cord cell line and its application

A technology of tissue cells and cell lines, applied in the field of cell culture, can solve the problems of slow growth, CyHV-2 reproduction, aggravating time costs, etc., achieve stable virulence, avoid repeated inoculation of cells, and reduce production costs

Active Publication Date: 2019-10-08
成都史纪生物制药有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004]CyHV-2 cannot continue to multiply in cells, so researchers who study CyHV-2 must inoculate cells repeatedly, which is time-consuming and labor-intensive
[0005] In addition, the culture temperature of commonly used fish cell lines cannot exceed 30°C, and their growth rate is very slow: under the condition of passage ratio 1:2, each generation The cultivation time is usually 10-14 days, which increases the time cost

Method used

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  • Carp spinal cord cell line and its application
  • Carp spinal cord cell line and its application
  • Carp spinal cord cell line and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 2

[0051] Example 2 Identification

[0052] The above-mentioned C8C37 F70 generation crucian carp spinal cord tissue cell line is further observed and identified (identification items include cryopreservation recovery experiment, chromosome analysis), and it is found that the cell line has the following biological characteristics:

[0053] (1) Morphology: The cell type was fibroblast-like cells, with many short synapses, which were judged to be glial cells according to the separation site.

[0054] (2) Growth characteristics: The passaged C8C37 cells began to adhere to the wall after 20 minutes of passage, and the healthy and living cells had completely adhered to the wall after about 5 hours. The population doubling rate is about 36h. The cells grow as dense multilayer cells, and the density effect is obvious. The higher the cell seeding density, the faster the proliferation rate.

[0055] (3) Stability: As of the date of application, the cells have been subcultured to the F75...

Embodiment 3

[0059] Embodiment 3 virus experiment

[0060] The application of the sensitive crucian carp spinal cord tissue cell line of CyHV-2, its step is as follows:

[0061] (1) Collection of tissue toxin samples: The tissue toxin suspension that was fatal due to CyHV-2 infection provided by Zhejiang Danshui Fisheries Research Institute was filtered through a 0.22 μm filter membrane to prepare a sterile tissue toxin suspension, and stored at -80°C after aliquoting. ℃.

[0062] (2) Proliferation of CyHV-2 on C8C37 cells: culture C8C37 cells T75 to about 80% monolayer, discard the medium, rinse with the preheated complete medium of No. ③ L15, and put the sterile tissue prepared above Inoculate 0.5ml of the virus suspension into a T75 cell flask, absorb for 1 hour at 24°C, and shake 3 times during this period to ensure uniform virus adsorption. After the adsorption was completed, 15ml of L15 maintenance solution with a concentration of 2% neonatal bovine serum was added to continue cult...

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Abstract

The invention provides a carp spinal cord cell line, which is named as a spinal cord cell line C8C37 of carassius auratus gibelio, which is preserved on the Chinese Culture Collection Center on November 15, 2018, and a preservation number is CCTCC NO: C2018204. The cell line can be rapidly propagated at 37 DEG C and the virus be can stably passed. The invention also provides a breeding method based on the spinal cord cell lines with similar properties. The cell line of the present invention can be well applied to the research of herpes simplex virus type 2 and the production of related vaccines, and has the advantages of low cost, low time consumption, and good effect.

Description

technical field [0001] The invention relates to the field of cell culture, in particular to a crucian carp spinal cord tissue cell line suitable for culturing carp herpesvirus type 2. Background technique [0002] In 1992, a large number of juvenile goldfish cultured in western Japan died, and the mortality rate reached almost 100%. The pathogen was identified as Herpesviral haemato poietic necrosis virus (HVHNV), because it was the second from Cyprinid herpesvirus 2 (Cyprinid herpesvirus 2, CyHV-2) is a herpes virus isolated from carps. From 2007 to 2008, some breeding farms in Yancheng, Jiangsu, my country found that the cultured crucian carp had surface hemorrhage and gill hemorrhage as the main symptoms, and the mortality rate was as high as 90%. After 2009, the disease showed a trend of large-scale outbreaks in successive years in the main crucian carp producing area. And with Yancheng, Jiangsu as the radiation point, it spread to Jiangxi, Hubei, Anhui, Zhejiang, Guan...

Claims

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Application Information

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IPC IPC(8): C12N5/079C12N7/00C12N7/02C12R1/91
CPCC12N5/0618C12N7/00C12N2710/16051Y02A40/81
Inventor 邢刚周涛岳丰雄崔静雯黄杰王洁清沈锦玉曹铮潘晓艺夏焱春姚嘉赟蔺凌云尹文林刘忆瀚
Owner 成都史纪生物制药有限公司
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