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H9N2 swine influenza virus and its kit

A swine influenza virus and kit technology, which is applied in the detection/examination of viruses/phages, microorganisms, microorganisms, etc., can solve the problems of inability to confirm and prevent swine influenza virus, transmission, etc.

Inactive Publication Date: 2013-09-25
SHANGHAI ANIMAL EPIDEMIC PREVENTION & CONTROL CENT
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] In order to solve the above-mentioned technical problems existing in the prior art, the present invention provides new H9N2 swine influenza virus and its test kit, and described this new H9N2 swine influenza virus and its test kit will solve the problems that cannot be confirmed in the prior art. and technical issues in preventing new swine flu viruses from spreading to pigs or humans from new swine flu outbreaks

Method used

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  • H9N2 swine influenza virus and its kit
  • H9N2 swine influenza virus and its kit
  • H9N2 swine influenza virus and its kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0053] Example 1 Acquisition of virus

[0054] It was obtained from a slaughterhouse in Shanghai in 2009, and the collected samples were pig lungs. Thoroughly grind the lung samples identified as positive, add antibiotic-containing PBS to make a tissue suspension with a volume fraction of 20%, freeze-thaw three times, centrifuge at 3000g for 20min, take the supernatant, and add penicillin at a final concentration of 1000IU / mL And the final concentration is 1000mg / mL streptomycin, after 1h at 37℃, used for chicken embryo inoculation. The processed samples were inoculated with 9-day-11-day-old SPF chicken embryos through the allantoic cavity at 0.2mL / embryo. Each sample was inoculated with 5 embryos and incubated in an incubator at 35℃~37℃. After 18 hours The death of chicken embryos was observed every 8h. The dead embryos after 18h and the allantoic fluid of chicken embryos still alive in 96h were aseptically collected, and the hemagglutination activity was measured. The virus i...

Embodiment 2

[0055] Example 2 Complete gene sequencing of two H9N2 swine influenza viruses

[0056] 2.1 In 2009, a pig lung sample was obtained from a slaughterhouse in Shanghai, and two H9N2 swine influenza viruses were isolated and identified: A / Swine / Henan / Y1 / 09; A / Swine / Shanghai / Y1 / 09. The H9 subtype serum used for virus identification was purchased from Harbin Veterinary Research Institute of Chinese Academy of Agricultural Sciences. The specific primers designed to amplify all 8 gene segments of the H9N2 subtype swine influenza virus were synthesized by Bao Biological (Dalian) Engineering Company.

[0057] HA gene upstream primer P1: 5’-CAGGGGAATTTCACAACCAGTCAAA-3’;

[0058] Downstream primer of HA gene P2: 5'-GCCAGGGTGTTTTTGCCAATTATATAC-3'.

[0059] M gene upstream primer P1: 5’-AAGATGAGCCTTCTAACCGAGGTC-3’;

[0060] M gene downstream primer P2: 5'-CGGGGTAGTTTTTTACTCCAGCTCTATG-3'.

[0061] NA gene upstream primer P1: 5’-AGCAGGAGTGAAAATGAATCCA-3’;

[0062] Downstream primer of NA gene P2: 5'-AA...

Embodiment 3

[0125] Example 3 Comparison of the whole sequence of HA and NA and analysis of some key sites

[0126] The homology of HA genes of the two swine influenza viruses is 96.6%, and the homology of NA genes is 98.6%. The analysis of http: / / www.cbs.dtu.dk / services / NetNGlyc / shows that the HA1 molecules of these two strains have 6 common glycosylation sites, which are 11, 64, 123, 200, At positions 280, 287 and 295, there are 2 potential glycosylation sites on the HA2 molecule, 474 and 533. Table 1 shows the characteristic amino acid analysis of the HA cleavage site and receptor binding site of these two strains and the 8 published H9N2 subtype swine influenza viruses. The sequence of the HA cleavage site indicates that they are all non-highly pathogenic Sex strain. Through blast, the HA gene fragment of Swine / Henan / Y1 / 09 has higher homology with the avian H9N2 subtype influenza virus isolated in 1999, while the NA gene fragment has a higher homology with the avian H9N2 subtype influe...

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Abstract

The invention belongs to the field of biotechnology and relates to a novel separated strain swine influenza virus. The invention discloses two separated H9N2 swine influenza viruses. According to the invention, it firstly proves that the two strains both belong to Ck / Bei series. The invention also discloses a kit for detecting the H9N2 swine influenza viruses. Through the kit provided by the invention, whether swine flu is caused by the genotype of H9N2 swine influenza viruses can be rapidly detected or eliminated so as to control the swine flu epidemic situation as soon as possible. In addition, by the use of the kit, bioinformation for a swine influenza virus monitoring system can be provided.

Description

Technical field [0001] The invention belongs to the field of biotechnology, and relates to isolated new swine influenza viruses, especially new H9N2 swine influenza viruses and kits thereof. Background technique [0002] Swine influenza (SI) is an important swine respiratory disease caused by swine influenza virus (SIV). It is prevalent all over the world and caused great economic losses to the pig industry. SIV belongs to the genus of influenza A virus of the Orthomyxoviridae family. The first H9N2 subtype avian influenza virus (AIV) was isolated from Guangdong Province in 1994 in my country. Since then, this subtype virus has spread across the country. . The H9N2 subtype AIV can not only infect poultry, but also pigs and humans. The occurrence of the 1997 Hong Kong flu incident that shocked the world made people realize that AIV can directly infect people without the adaptation of an intermediate host to break through the interspecies barrier. In this incident, 18 people were ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/00C12Q1/70C12Q1/68C12R1/93
Inventor 刘佩红周锦萍葛菲菲刘健鞠厚斌杨德全
Owner SHANGHAI ANIMAL EPIDEMIC PREVENTION & CONTROL CENT
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