A kind of cultivating method for improving the virus titer of duck flavin virus

A culture method, duck flavivirus technology, applied in the directions of microorganism-based methods, viruses/phages, biochemical equipment and methods, etc., can solve the problems of slow primary virus culture process and insufficient virus titer, and achieve increased affinity. the effect of improving the infection efficiency

Active Publication Date: 2017-01-18
GUANGDONG HAID ANIMAL HUSBANDRY & VETERINARY RES INST +1
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  • Summary
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  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Many literature reports pointed out that the clinically isolated duck yellow virus can infect a variety of cells in vitro, such as VERO cells, BHK cells, etc., but the primary virus culture process is relatively slow, and the virus titer is not high enough. a problem to be solved

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0055] A kind of cultivation method that improves duck flavin virus virus titer, it comprises the following steps:

[0056] A. Preparation of DMEM culture solution: Dissolve 13.4g DMEM dry powder in 1000ml deionized water, stir until completely dissolved, adjust the pH value to 7.0, and use a 0.22um sterile filter membrane for sterilizing filtration, and place it in a refrigerator at 4°C stand-by;

[0057] B. After adding fetal calf serum accounting for 5% of the volume of DMEM culture medium in the above-mentioned DMEM solution, inoculate Vero cells and carry out monolayer culture;

[0058] C. After the monolayer Vero cells are cultured, discard the DMEM culture medium in the culture container, and wash with sterile deionized water for 3 times;

[0059] D, discard the sterile deionized water, then add affinity agent and the DMEM culture solution described in step A in the culture container, wherein the volume consumption of the affinity agent is 1% of the volume consumption ...

Embodiment 2

[0069] A kind of cultivation method that improves duck flavin virus virus titer, it comprises the following steps:

[0070] A. Preparation of DMEM culture solution: Dissolve 13.4g DMEM dry powder in 1000ml deionized water, stir until completely dissolved, adjust the pH value to 6.8, and use a 0.22um sterile filter membrane for sterilizing filtration, and place it in a refrigerator at 4°C stand-by;

[0071] B. After adding fetal calf serum accounting for 8% of the volume of DMEM culture medium to the above-mentioned DMEM solution, inoculate 293 cells for monolayer culture;

[0072] C. After the monolayer of 293 cells is cultured, discard the DMEM culture medium in the culture container, and wash with sterile PBS buffer 3 times;

[0073] D, discard the sterile PBS buffer, then add the affinity agent and the DMEM culture solution described in step A to the culture container, wherein the volume consumption of the affinity agent is 0.5% of the volume consumption of the DMEM cultur...

Embodiment 3

[0083] A kind of cultivation method that improves duck flavin virus virus titer, it comprises the following steps:

[0084] A. Preparation of DMEM culture solution: Dissolve 13.4g DMEM dry powder in 1000ml deionized water, stir until completely dissolved, adjust the pH value to 7.4, and use a 0.22um sterile filter membrane for sterilizing filtration, and place it in a refrigerator at 4°C stand-by;

[0085] B. After adding fetal calf serum accounting for 6% of the volume of DMEM culture medium in the above-mentioned DMEM solution, inoculate ST cells and carry out monolayer culture;

[0086] C. After the monolayer ST cells are cultured, discard the DMEM culture medium in the culture container, and wash twice with sterile serum-free DMEM culture medium;

[0087] D, discard the sterile serum-free DMEM culture solution, then add the DMEM culture solution described in the affinity agent and step A in the culture container, wherein the volume consumption of the affinity agent is 0.8...

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Abstract

The invention discloses a culture method for increasing the virus titer of duck flavivirus, and belongs to the field of microbial virus technologies. The culture method mainly comprises the following steps: preparing a DMEM culture solution; after fetal calf serums are added, inoculating cells for carrying out monolayer culture; after an affinity absorber and the DMEM culture solution are added, further culturing for 1-30 min, and inoculating duck flavivirus for 2-5 days, so that duck flavivirus liquid with a high virus titer can be obtained. The culture method disclosed by the invention increases the contact affinity of viruses and cells mainly by changing a microenvironment between the duck flavivirus and the cells, thereby greatly increasing the virus titer of the duck flavivirus.

Description

technical field [0001] The invention relates to the technical field of microbial viruses, in particular to a culture method for increasing the titer of duck yellow virus. Background technique [0002] Duck yellow virus (Duck Tembusu virus) is a virus that can seriously reduce the egg production of laying ducks. It can be reduced to zero, and the feed intake will also decrease at the same time, and some of them will die. Pathological dissection showed serious lesions such as ovarian hyperemia, swelling, necrosis, and atrophy. Since the disease was first discovered in China in 2010, it has caused a large-scale reduction in the production of egg ducks in Shandong, Hunan, Guangdong and other major duck breeding provinces. At present, many research institutions have clinically isolated duck yellow virus, and carried out cell infection tests in vitro, trying to propagate the virus through in vitro culture in order to be used in the development of duck yellow virus vaccine. Many...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N7/00C12R1/93
Inventor 冯晓声王贵平贾爱卿王伟
Owner GUANGDONG HAID ANIMAL HUSBANDRY & VETERINARY RES INST
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