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A largemouth bass brain cell line and its application

A technology for largemouth bass and brain cells, applied in animal cells, cell dissociation methods, nervous system cells, etc., can solve problems such as large economic losses and poor drug effects, and achieve reduced dosage, small errors, and good repeatability Effect

Active Publication Date: 2022-07-26
HUAZHONG AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

In recent years, California perch rhabdovirus disease has broken out in large numbers during the cultivation of California perch seedlings. When it is severe, 80% of the ponds are infected, and the mortality rate can be as high as 90% in a short period of time. The drug effect is not good, and the economic loss is huge.

Method used

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  • A largemouth bass brain cell line and its application
  • A largemouth bass brain cell line and its application
  • A largemouth bass brain cell line and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Example 1 Separation and culture of largemouth bass brain cells

[0037] 1. Experimental reagents and materials

[0038] (1) Experimental animals: It was repeatedly verified by molecular methods that healthy largemouth bass that were not infected with aquatic animal viruses, about 10 cm in size, came from the largemouth bass seedling farm in Zhijiang City, Hubei Province, and were reared in the greenhouse at the base of the School of Fisheries of Huazhong Agricultural University. in a breeding tank.

[0039] (2) Experimental equipment: scalpel, ophthalmic scissors, ophthalmic forceps, 70 μm cell mesh screen, syringe, alcohol cotton and gauze, etc.

[0040] (3) Experimental reagents: AIM medium, liquid medium M199, trypsin-EDTA, penicillin / streptomycin 10000U / mL, fetal bovine serum (FBS) were purchased from Gibco; amphotericin B, gentamicin , Colchicine were purchased from Biosharp Company; dimethyl sulfoxide (DMSO) was purchased from Sigma Company, fibroblast growth fac...

Embodiment 2

[0060] Embodiment 2 The optimum culture environment, growth curve and chromosome analysis of largemouth bass brain cells

[0061] 1. Optimum culture temperature of largemouth bass brain cell line MSBr

[0062] Select four different culture temperatures of 16, 22, 28, and 37 °C, use M199 medium supplemented with 10% FBS, and adjust the cell density to 1 × 10 5 cells / mL, and the cell suspension was seeded in a 12-well plate at a volume of 500 μL / well and placed in 4 incubators with different culture temperatures. Take out 3 wells of cells from each experimental group every 1 d, collect and count cells by Trypsin-EDTA digestion, co-culture for 5 d, count 5 times in a row, draw their growth curves, and the cell growth curves at different culture temperatures are as follows: figure 2 shown. The results showed that the largemouth bass brain cell line MSBr could grow at four temperatures, of which 28°C was the optimal culture temperature for MSBr cells.

[0063] 2. Optimal fetal ...

Embodiment 3

[0069] Example 3 Sensitivity test of largemouth bass brain cell line MSBr to different viruses

[0070] 1. Sensitivity test of largemouth bass brain cell line MSBr to different aquatic animal viruses

[0071] The cells were digested with Trypsin-EDTA and adjusted to a cell density of 1 × 10 51 mL / well inoculated into 6-well plates. When the cells just covered the well plate, after washing twice with PBS buffer, 1 MOI of different viruses (largemouth bass rhabdovirus MSRV, carp spring viremia virus SVCV, grass carp reovirus GCRV, frog iris virus RGV , red-spotted grouper nerve necrosis virus RGNNV, infectious hematopoietic organ necrosis virus IHNV, infectious spleen and kidney necrosis virus ISKNV) 7 kinds of viruses were inoculated into the largemouth bass brain cell line MSBr of the present invention, and the inoculation amount was 200 μL. After inoculation, the CPE production of the cells was observed, and the cells without virus inoculation were used as the control (NC),...

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Abstract

The invention discloses a largemouth bass brain cell line and an application thereof. The largemouth bass brain cell line has been deposited in the China Center for Type Culture Collection on March 25, 2021, and the deposit number is CCTCC NO: C202166. The largemouth bass brain cell line MSBr described in the present invention is stable in passage, has been successfully passaged for more than 65 generations and has a good growth state, and the cell line has a wide range of applications, not only for virus isolation and identification, pathogenic function research, and environmental pollutant detection. , gene screening and functional analysis and fish tumor research, etc., and can also be used for monitoring, diagnosis and epidemiological investigation of aquatic animal viral diseases, development of diagnostic kits and vaccines for related viral diseases, etc. application value and socio-economic value.

Description

technical field [0001] The invention belongs to the technical field of aquatic biological cells and aquaculture disease prevention and control, and in particular relates to a largemouth bass brain cell line and applications thereof. Background technique [0002] Cell culture research in aquatic animals (mainly fish) was later than mammals and began in the 1960s. In 1962, Woft and Quimby established the world's first fish cell line, the rainbow trout gonadal cell line. Since then, fish cell culture research has been very rapid, and 275 fish cell lines have been reported worldwide. The culture of fish cells in my country began in the late 1970s. The grass carp rostral tissue diploid cell line established by Zhang Nianci and others was the first fish cell line in my country. There are more than 70 fish cell lines established so far, most of which are from marine fish. As a biologically active in vitro material, cell lines have the incomparable advantages of live fish: the cu...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N5/079C12Q1/02C12R1/91
CPCC12N5/0618G01N33/5058C12N2509/00C12N2509/10C12N2503/02G01N2500/10
Inventor 袁军法戴彩姣余利李莉娟
Owner HUAZHONG AGRI UNIV
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