Primer pair composition and application thereof for identifying or assisting in identifying h6n1 subtype avian influenza virus

A technology for auxiliary identification of avian influenza virus, applied in the direction of recombinant DNA technology, microbial measurement/testing, biochemical equipment and methods, etc., can solve the problems of long detection cycle and achieve high sensitivity and high specificity

Active Publication Date: 2016-06-29
GUANGXI VETERINARY RES INST
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] Virus isolation and identification is a classic influenza detection method, that is, the sample is first inoculated with SPF chicken embryo proliferation virus for 2-5 days, and then the chicken embryo allantoic fluid is collected for hemagglutination test and hemagglutination inhibition test. The results are accurate and reliable, but there are detection Disadvantages of long cycle
Enzyme-linked immunosorbent assay can detect antigens of avian influenza, but requires specific standard positive sera

Method used

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  • Primer pair composition and application thereof for identifying or assisting in identifying h6n1 subtype avian influenza virus
  • Primer pair composition and application thereof for identifying or assisting in identifying h6n1 subtype avian influenza virus
  • Primer pair composition and application thereof for identifying or assisting in identifying h6n1 subtype avian influenza virus

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0067] Embodiment 1, preparation identification or auxiliary identification primer pair composition of H6N1 subtype avian influenza virus

[0068] A primer pair composition for identification or auxiliary identification of H6N1 subtype avian influenza virus (AIV), consisting of a PCR primer pair named H6 and a PCR primer pair named N1; said H6 consists of the sequence shown in SEQIDNo.1 Composed of single-stranded DNA (H6-F) and single-stranded DNA (H6-R) shown in SEQIDNo.2, a band with a size of 447bp can be amplified from the avian influenza virus of the H6 subtype; the N1 is composed of the sequence The single-stranded DNA (N1-F) shown in SEQIDNo.3 and the single-stranded DNA (N1-R) shown in SEQIDNo.4 in the list consist of a single-stranded DNA (N1-R) that can be amplified from the avian influenza virus of N1 subtype with a size of 325bp Bands.

[0069] In the primer pair composition for identifying or assisting in identifying the H6N1 subtype avian influenza virus, the H...

Embodiment 2

[0070] The identification of embodiment 2, embodiment 1 or the primer that assists the identification of H6N1 subtype avian influenza virus to the sensitivity experiment of composition

[0071] According to the instructions of the MiniBESTViralRNA / DNAExtraction kit, the total RNA of the H6N1 subtype avian influenza virus (referred to as H6N1RNA) was extracted from the H6N1 subtype avian influenza virus at a concentration of 10 ng / μl.

[0072] Use HoffmannE, StechJ, GuanY, etal.Universalprimersetforthefull-lengthamplificationofallinfluenzaAviruses[J].ArchVirol,2001,146:2275-2289 respectively in the literatureHoffmannE, StechJ,GuanY,etal.Universalprimersetforthefull-lengthamplificationofallinfluenzaAviruses[J].ArchVirol,2001,146:2275-2289The RT-PCR amplification is carried out with H6N1RNA as a template. The reaction system is as follows: 2 ×1StepBuffer12.5μL, PrimeScript1StepEnzymeMix1μL, HA or NA gene upstream primer (25μM) 0.5μL, HA or NA gene downstream primer (25μM) 0.5μL, H...

Embodiment 3

[0081] The identification of embodiment 3, the specificity experiment of the primer of embodiment 1 or the primer pair composition of auxiliary identification avian influenza virus

[0082] 1. Preparation of test samples

[0083] The viruses used in the experiment are H6N1 subtype avian influenza virus, H6N2 subtype avian influenza virus, H6N5 subtype avian influenza virus, H6N6 subtype avian influenza virus, H6N8 subtype avian influenza virus, H1N1 subtype avian influenza virus, H2N3 subtype avian influenza virus, H3N2 subtype avian influenza virus, H4N5 subtype avian influenza virus, H5N1 subtype avian influenza virus, H7N2 subtype avian influenza virus, H8N4 subtype avian influenza virus, H9N2 subtype avian influenza virus, H10N3 Avian influenza virus subtype, H11N9 subtype avian influenza virus, H12N5 subtype avian influenza virus, H13N5 subtype avian influenza virus and H15N8 subtype avian influenza virus, Newcastle disease virus (NDV), infectious bronchitis virus (IBV) a...

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Abstract

The invention discloses a primer pair composition for identification or assisted identification of H6N1 subtype of avian influenza virus and application of the primer pair composition. The primer pair composition for identification or assisted identification of H6 and / or N1 subtype of avian influenza virus is composed of two primer pairs which are independently packaged, namely a polymerase chain reactor (PCR) primer pair named as H6 and a PCR primer pair named as N1; the H6 is composed of two single-stranded DNA as shown in SEQ ID NO. 1 and SEQ ID NO. 2; and the N1 is composed of two single-stranded DNA as shown in SEQ ID NO. 3 and SEQ ID NO. 4.

Description

technical field [0001] The invention relates to a primer pair composition and application thereof for identifying or assisting in identifying H6N1 subtype avian influenza virus in the field of biomedicine. Background technique [0002] Avian Influenza Virus (AIV) is divided into different subtypes according to the antigenic differences of surface glycoprotein (Hemagglutinin, HA) and neuraminidase (Neuraminidase, NA). So far, 16 HA subtypes (H1-H16) have been found. ) and 9 NA subtypes (N1-N9), the combination of HA and NA can produce at least 135 subtypes, among which, some combinations of H5 and H7 subtypes (H5N1, H7N7 and H7N9, etc.) are called highly pathogenic Sexual AIV, which can infect humans and cause death. Although low-pathogenic AIV infection has no symptoms or shows mild and mild atypical symptoms, it is not a threat to the health of livestock and humans, but when different subtypes of low-pathogenic AIV strains mix to infect livestock and poultry , can exchang...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/70C12Q1/68C12N15/11
CPCC12Q1/701C12Q2600/16
Inventor 谢芝勋罗思思谢志勤邓显文刘加波黄莉黄娇玲曾婷婷
Owner GUANGXI VETERINARY RES INST
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