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Newcastle disease virus marker vaccine strain of genotype ⅶ and its application

A vaccine strain and labeling technology, applied in antiviral agents, viruses/phages, viral antigen components, etc., can solve the problems of reducing the virulent carrying capacity and infection rate of NDV, achieve good immune protection effect, inhibit detoxification, and be widely used effect of value

Active Publication Date: 2021-01-26
CHINA AGRI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Traditional vaccines cannot provide ideal immune protection against the current virulent strains of NDV. Domestic and foreign studies have proved that the use of gene-VII vaccines consistent with the epidemic strains can effectively reduce the carrying amount and infection of virulent NDV in immunized chickens. rate, therefore, it is necessary to develop new alternative vaccines to deal with the prevalence of Newcastle disease

Method used

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  • Newcastle disease virus marker vaccine strain of genotype ⅶ and its application
  • Newcastle disease virus marker vaccine strain of genotype ⅶ and its application
  • Newcastle disease virus marker vaccine strain of genotype ⅶ and its application

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0046] Example 1 Rescue of aSG10 strain

[0047] 1. Construction of full-length cDNA of SG10 strain genome

[0048] (1) Purification of the virus In order to obtain a single virus clone, the SG10 strain (disclosed in the literature Liu MM, et al. Generation by reverse genetics of an effective attenuated Newcastle disease virus vaccine based on a prevalent highly virulent Chinese train) .BIOTECHNOL LETT 2015 2015-06-01; 37(6):1287-96.) was purified, the detailed steps are as follows: the virus liquid was diluted 10 times, and after dilution, 100 μL of each titer virus liquid was inoculated with SPF For each dilution, 4 chicken embryos were collected, and the chicken embryos that died within 24 hours were discarded, and the allantoic fluid of the chicken embryos was harvested 4 days later to measure the HA activity. Select the allantoic fluid with the highest dilution factor of HA activity to do the same multiple dilution and inoculation. After the virus was continuously passa...

Embodiment 2

[0085] Example 2 Rescue of aSG10-mHN labeled vaccine strain

[0086] 1. Test method

[0087] 1.1 Construction of full-length cDNA of aSG10-mHN strain genome

[0088]The inserted molecular tag (CACCACCACCAC) is located at fragment D of the full-length cDNA plasmid pOK-aSG10. Design primers (Table 7), insert a molecular tag on the D fragment of pOK-aSG10 (the 5' non-coding region of the HN gene, located at position 8140 in the whole genome) by fusion PCR, and combine the mutated fragment mD with the full-length The D fragment of plasmid pOK-aSG10 was replaced to construct the full-length genome plasmid pOK-aSG10-mHN containing molecular tags. See the construction diagram image 3 .

[0089] Table 7 Primers used to amplify mD fragments

[0090]

[0091] Note: The bold part is the stop codon of HN gene ORF, and the underlined part is the insertion mutation part.

[0092] The fragment after PCR cloning with primers D-U and mHN-L was named mD1, and the fragment after PCR cl...

Embodiment 3

[0109] Example 3 aSG10-mHN labeled vaccine strain to SPF chicken safety test and challenge protection test

[0110] 1 Test method

[0111] 1.1 Safety test of aSG10-mHN labeled vaccine strain on SPF chickens

[0112] In order to measure the safety of the aSG10-mHN labeled vaccine strain rescued by Example 2 of the present invention on SPF chickens, the 2-week-old SPF chickens reared in isolators were randomly divided into 4 groups, 19 in each group. The first group is the aSG10-mHN immunization group, the second group is the aSG10 immunization group, and each chicken is treated with 10 6.0 EID 50 Dosage: Inoculate aSG10-mHN or aSG10 by nasal drop or eye drop respectively. The third group was the blank control group, which was replaced by an equal volume of normal saline during immunization. On the 3rd and 7th days after inoculation, 3 chickens in each group were randomly killed, the gross pathological changes were recorded, and the spleen, thymus, bursa of Fabricius, Harder...

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Abstract

The present invention provides a gene type VII NDV labeled vaccine strain aSG10-mHN, the labeled vaccine strain is an attenuated strain aSG10 with the nucleotide sequence shown in SEQ ID NO.1 in the 5' non-coding region of the HN gene. The virulence of the marked vaccine strain constructed by the invention is obviously reduced and heredity is stable. The results of the immune protection test showed that the marked vaccine strain had a good immune protection effect against NDV, and could effectively inhibit the shedding of the virus, and could be used to prevent the current epidemic genotype VII Newcastle disease virus. Furthermore, the PCR method can be used to identify wild virus-infected animals and vaccine-immunized animals, which is of great significance to the monitoring, diagnosis, control and purification of ND, and has a wide range of application values.

Description

technical field [0001] The invention belongs to the field of veterinary biological products, in particular to a gene type VII NDV labeled vaccine strain aSG10-mHN and its application. Background technique [0002] Newcastle Disease (ND) is caused by Newcastle Disease Virus (NDV), and is an acute, febrile, highly contagious infectious disease characterized by damage to the digestive tract and respiratory tract of poultry. It spreads fast, has a high mortality rate, and is seriously harmful, causing huge economic losses to the poultry industry of various countries every year. Vaccine immunization is an important means to prevent Newcastle disease, but most of the vaccine strains currently used in China are genotype I or II, which are significantly different from the current prevailing genotype VII strains. Although traditional vaccines can induce a higher level of immune antibodies and resist Newcastle disease virus infection to a certain extent, they cannot completely preven...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N7/01C12N15/86A61K39/17A61P31/14C12Q1/70C12Q1/6869C12N15/11C12R1/93
Inventor 张国中赵静靳继惠邵梦瑜
Owner CHINA AGRI UNIV
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