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Method for producing porcine reproductive and respiratory syndrome viruses

A technology for respiratory syndrome and production method, applied in the field of virus culture, can solve the problems of long time required for virus liquid harvesting, high labor intensity of workers, hidden dangers of vaccine safety, etc., so as to save freezing and thawing time, reduce labor intensity, and reduce allergies source effect

Active Publication Date: 2010-06-23
WENS FOOD GRP CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The current culture of porcine blue ear disease virus (PRRSV) leaves a large amount of bovine serum and other allergens in the crude antigen, which brings certain safety hazards to the use of vaccines
In addition, it takes a long time to harvest the virus liquid and the procedures are relatively cumbersome, resulting in high labor intensity for workers

Method used

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  • Method for producing porcine reproductive and respiratory syndrome viruses
  • Method for producing porcine reproductive and respiratory syndrome viruses
  • Method for producing porcine reproductive and respiratory syndrome viruses

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0023] Test group: In 1700mL spinner bottle and 15000mL spinner bottle, add DMEM (high sugar type, containing 4500mg / L D-glucose, L-glutamine, and 110mg / L sodium pyruvate, without sodium bicarbonate) +10 %newborn bovine serum+1% double antibody (penicillin and streptomycin) was used as culture medium to cultivate Marc-145 cells; after the cells in the spinner bottle grew into a single layer, the cell growth solution in the bottle was discarded, and PRRSV was inoculated at a multiplicity of infection of 0.01. After adsorption at 37°C for 1 hour, DMEM culture solution was added as a maintenance solution, and placed on a rotary bottle machine in a constant temperature room at 37°C for 75 hours of rotation cultivation. When 75%-80% CPE appeared in Marc-145 cells, the cell supernatant was collected to harvest the virus. Determination of half tissue culture infectious dose (TCID) in 96-well plate 50 ).

[0024] At the same time, a control group was set up, the maintenance solution...

Embodiment 2

[0039] Test group: In 1700mL spinner bottle and 15000mL spinner bottle, add DMEM (high sugar type, containing 4500mg / L D-glucose, L-glutamine, and 110mg / L sodium pyruvate, without sodium bicarbonate) +10 %newborn bovine serum+1% double antibody (penicillin and streptomycin) was used as culture medium to cultivate Marc-145 cells; after the cells in the spinner bottle grew into a single layer, the cell growth solution in the bottle was discarded, and PRRSV was inoculated at a multiplicity of infection of 0.01. Adsorbed at 37°C for 1 hour, added DMEM culture medium as a maintenance solution, and placed in a 37°C thermostatic chamber rotary bottle machine for 78 hours of rotary culture. When 75%-80% CPE appeared in Marc-145 cells, the cell supernatant was collected to obtain PRRSV.

[0040] Control group: the same as the experimental group, but after collecting the supernatant, put it in the freezer and freeze and thaw it repeatedly 3 times to obtain PRRSV.

[0041] When the two ...

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Abstract

The invention discloses a method for producing and harvesting porcine reproductive and respiratory syndrome viruses (PRRSV), which comprises the following steps: culturing Marc-145 cells in a spinner bottle; after the cells grow into a monolayer, removing a growth-promoting media; inoculating the PRRSV according to 0.01 infection multiplicity (MOI: multiplicity of infection); absorbing for 1 hour at a temperature of 37 DEG C; adding DMEM serving as a maintenance media to carry out expanding propagation of viruses; arranging the spinner bottle on a rotary machine of a 37 DEG C thermostatic chamber to carry out rotation culture for 72 to 84 hours; and when 75 to 80 percent of cytopathic effect occurs, directly collecting the supernatant to obtain the PRRSV. The invention reduces allergen in the rough antigens and improves the vaccine quality. And the time that the Marc-145 cells have CPE is consilient with that before the process is improved, but the virus harvesting time is shortened.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a method for culturing viruses. Background technique [0002] Since its discovery in the United States in 1987, reproductive and respiratory syndrome (PRRS) has spread to almost all countries and regions with developed swine industry in the world. The clinical symptoms are mainly characterized by reproductive disorders in sows and respiratory diseases in piglets. Highly pathogenic porcine blue ear disease is an acute and highly pathogenic disease caused by a mutant strain of porcine reproductive and respiratory syndrome (commonly known as blue ear disease), which has caused huge economic losses to the world's pig industry . Effective vaccine immunization is an important measure to control the disease. In order to prepare a large number of virus antigens, enough virus must be obtained. Viruses are usually propagated by multiplying on cells. Porcine Reproductive and Resp...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/00C12N7/02
Inventor 邓雨修宋延华杨明柳李春梅文传洪于琳忠
Owner WENS FOOD GRP CO LTD
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