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MRT-PCR (multiple reverse transcription-polymerase chain reaction) kit for discriminating and detecting important avian economic animal viruses and application of mRT-PCR kit

A kit and bird flu virus technology, applied in the field of mRT-PCR kits, can solve the problems of time-consuming and labor-consuming, disease prevention and control detection, insufficient biosafety of breeding enterprises, difficulties in the isolation and identification of poultry respiratory pathogens, etc., and achieve high sensitivity , time-saving and labor-saving material reagents, and the effect of reducing the loss of aquaculture

Active Publication Date: 2015-07-29
山西隆克尔生物制药有限公司 +2
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although there are attenuated and inactivated vaccines on the market to control the epidemic, it is still not enough for disease prevention and control detection and the biosecurity of breeding enterprises
Virus isolation, identification and serological tests are usually used in the diagnosis of virus infection, but these methods are time-consuming and labor-intensive, especially the large-scale immunization of vaccines makes the isolation and identification of poultry respiratory pathogens extremely difficult

Method used

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  • MRT-PCR (multiple reverse transcription-polymerase chain reaction) kit for discriminating and detecting important avian economic animal viruses and application of mRT-PCR kit
  • MRT-PCR (multiple reverse transcription-polymerase chain reaction) kit for discriminating and detecting important avian economic animal viruses and application of mRT-PCR kit
  • MRT-PCR (multiple reverse transcription-polymerase chain reaction) kit for discriminating and detecting important avian economic animal viruses and application of mRT-PCR kit

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0047] Embodiment 1, preparation and assembly of kit

[0048] 1. Design and synthesis of primers

[0049] Referring to the published sequences of the popular strains in China in the gene bank, and through software comparison and optimization, design and synthesis were used to detect avian influenza virus (Avian Influenza virus, AIV) M gene, avian influenza virus H7, H9, H5 subtypes, Newcastle disease Newcastle disease virus (NDV), infectious bronchitis virus (IBV), infectious laryngotracheitis virus (ILTV), infectious bursal disease virus (IBDV) As well as the primer pair for Egg drop syndrome virus (Egg drop syndrome virus 76, EDS 76), the specific primer sequences are shown in Table 1 below, and each primer was diluted at 100 pmol / μL and stored at -20°C for later use.

[0050] Table 1 The primer sequences involved in multiple reverse transcription polymerase chain reaction (mRT-PCR)

[0051]

[0052]

[0053] Wherein the mixed coding site means: Y=C / T; R=A / G; W=A / T; ...

Embodiment 2

[0056] Example 2 Application of the mRT-PCR kit of the present invention in distinguishing and detecting poultry important economic viruses

[0057] 1. Viruses and samples

[0058] The following viruses were used to standardize multiplex RT-PCR:

[0059] H7N9 virus isolated

[0060] 1.A / Chicken / Weixian / China 2013-4 / 03(H7N9)

[0061] 2.A / Chicken / Weixian / China / 2013-5 / 04(H7N9)

[0062] 3.A / Chicken / Weixian / China / 2014-4 / 04(H7N9)

[0063] H9N2 Virus Isolation

[0064] 1.A / Chicken / / Xingtai / China / 2014-6 / 01(H9N2)

[0065] 2.A / Chicken / Xingtai / China / 2013-1 / 05(H9N2)

[0066] 3.A / Chicken / Weixian / China / 2013-1 / 06(H9N2)

[0067] H5N2 Virus Isolation

[0068] 1.A / Chicken / Weixian / China / 2013 / 03(H5N2)

[0069] 2.A / Chicken / Xingtai / China / 2013 / 04(H5N2)

[0070] 3.A / Chicken / Xingtai / China / 2013 / 06(H5N2)

[0071] The above strains were isolated from 9-day-old chicken embryos, identified, passed to 3 generations, and preserved in freeze-drying.

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Abstract

The invention discloses an mRT-PCR (multiple reverse transcription-polymerase chain reaction) kit for discriminating and detecting important avian economic animal viruses and an application of the mRT-PCR kit. The kit comprises primer pairs for detecting an AIV (avian influenza virus) M gene, AIV H7,H9 and H5 subtypes, an NDV (newcastle disease virus), an ILTV (infectious laryngotracheitis virus), an IBDV (infectious bursal disease virus) and an EDS (egg drop syndrome) virus. The mRT-PCR kit has the characteristics of high sensitivity, high specificity and the like when used for detecting AIVs and other common respiratory viruses, can be used for immediately detecting and discriminating important and popular subtypes of the AIVs and five avian respiratory pathogens most popular in China and has a great significance in zoonosis control, monitoring, prevention and control of avian mixed infection, biosafety of farms, reduction of loss of the breeding industry, food safety, quarantine and the like.

Description

technical field [0001] The invention relates to a kit for detecting various poultry viruses, in particular to an mRT-PCR kit for differentially detecting important economic viruses of poultry and its application. The invention belongs to the technical field of virus detection. Background technique [0002] Avian influenza (Avian influenza, AI) is a globally transmitted disease, caused by avian influenza viruses of the Orthomyxoviridae family. According to the classification of the virus surface hemagglutinin, 18 H subtypes have been found. 10 subtypes were found. Because the two proteins are highly variable, there are many different subtypes of avian influenza virus in nature. [0003] Many avian influenza viruses settle in the respiratory tract and digestive tract of birds and cause mild symptoms or no symptoms, but highly pathogenic avian influenza H5, H7, H9 subtype viruses can infect humans and cause disease. H5, H7, H9 avian influenza is prevalent in poultry, and H5, ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/68C12R1/93
CPCC12Q1/686C12Q1/701C12Q2600/16C12Q2537/143C12Q2521/107
Inventor 张澍吕宏亮
Owner 山西隆克尔生物制药有限公司
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