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Porcine epidemic diarrhea virus and separation culture method

A porcine epidemic diarrhea, isolation and cultivation technology, applied in the field of new porcine epidemic diarrhea virus and its isolation and cultivation, can solve the problems of unsuccessful attempts, difficult propagation of PEDV, and difficulty in isolation of PEDV

Inactive Publication Date: 2017-02-15
BEIJING DAWEIJIA BIOTECH SHARE CO LTD
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  • Abstract
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AI Technical Summary

Problems solved by technology

[0004] PEDV is difficult to proliferate in in vitro cultured cell lines, making the isolation of PEDV difficult
Since PED was first reported in the United Kingdom and Belgium in 1971, Japan, South Korea, Canada, China and other countries have successively reported the occurrence of the disease. Many scholars have used PK-15 cells (pig kidney cells), BHK cells (hamster kidney cells) Cells), bovine testicular cells, pig testicular cells, etc. to isolate and culture PEDV in vitro, but many years of attempts have been unsuccessful

Method used

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  • Porcine epidemic diarrhea virus and separation culture method
  • Porcine epidemic diarrhea virus and separation culture method
  • Porcine epidemic diarrhea virus and separation culture method

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Experimental program
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Embodiment 1

[0025] Embodiment 1: the acquisition of PEDV / CH / BJ / 2014 strain

[0026] 1. Collection and processing of disease materials

[0027] Collect 6 samples of feces and intestinal tissue from piglets with suspected viral diarrhea in a pig farm in Beijing. After the samples are numbered, add physiological saline at a mass volume ratio of 1:5, homogenize the intestinal tissue, shake and mix the fecal samples, and freeze and thaw repeatedly Centrifuge 3 times at 8000 rpm / min (rotation / minute) for 10 min, take the supernatant, first filter it with a filter with a pore size of 0.48 μm, and then use a filter with a pore size of 0.22 μm to filter and sterilize. Store in 80°C refrigerator for later use.

[0028] 2. RT-PCR (reverse transcription polymerase chain reaction) detection of disease materials

[0029] Take 250 μL of each of the above treatment solutions numbered 1-6, and use the Total RNA Kit II kit produced by OMEGA to extract RNA. The extracted RNA was reverse-transcribed: 6 μL...

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Abstract

The present invention discloses a strain of porcine epidemic diarrhea virus and a separation culture method. According to the present invention, the virus is Coronaviridae Coronavirus porcine epidemic diarrhea virus, is named PEDV / CH / BJ / 2014, and has the preservation number of CGMCC No.10111; the culture of the virus requires a blank DMEM culture liquid containing trypsin; and the porcine epidemic diarrhea virus has good immunogenicity, is the current domestic epidemic variant, can be used for preparing PEDV diagnosis kit, PEDV attenuated vaccines and inactivated vaccines, and enriches the existing PEDV vaccine resources.

Description

technical field [0001] The invention relates to the field of microbes, in particular to a new strain of porcine epidemic diarrhea virus and a method for its isolation and cultivation. Background technique [0002] Porcine Epidemic Diarrhea Virus (PEDV) is a member of the Coronaviridae family of Coronaviridae, which can cause intestinal infectious diseases of pigs characterized by vomiting, diarrhea and dehydration. Pigs of all ages are susceptible, but suckling piglets are the most affected, with mortality rates as high as 100%. In recent years, the outbreak of PED in many provinces in my country has caused serious economic losses to the pig industry. [0003] The prevention and control of porcine epidemic diarrhea disease is mainly based on vaccination. At present, there is no specific drug to treat this disease, and the vaccine used in China is mainly PEDV CV777 strain. This strain was isolated earlier. Compared with the new mutant strain Lack of extensive immunogenicity...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/00C12R1/93
Inventor 张美美李爽张志刚董建辉张晓杰闫雷张杏金忠辉
Owner BEIJING DAWEIJIA BIOTECH SHARE CO LTD
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