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Double LAMP detection kit for chicken infectious anemia and chicken parvovirus

A chicken infectious anemia, chicken parvovirus technology, applied in recombinant DNA technology, microbial assay/test, biochemical equipment and methods, etc., can solve the problem of chicken infectious anemia and chicken parvovirus double LAMP Visual detection kits and other issues

Active Publication Date: 2020-01-03
GUANGXI VETERINARY RES INST
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] At present, there are no relevant reports on the establishment of a double LAMP visual detection kit and method for chicken infectious anemia and chicken parvovirus at home and abroad

Method used

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  • Double LAMP detection kit for chicken infectious anemia and chicken parvovirus
  • Double LAMP detection kit for chicken infectious anemia and chicken parvovirus
  • Double LAMP detection kit for chicken infectious anemia and chicken parvovirus

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] Embodiment 1, the design of primer

[0028] According to the conserved sequences of the VP1 gene of CIAV (accession number: KY486154.1) and the NS1 gene of ChPV (accession number: KM254174.1) in GenBank, use the online software Primer Explorer V5 (http: / / primerexplorer.jp / lampv5e / index .html) to design LAMP primers. The primers were synthesized by Bao Biological Engineering (Dalian) Co., Ltd., and their specific sequences are shown in Table 1.

[0029] Table 1 LAMP primer sequence

[0030]

Embodiment 2

[0031] Example 2, Application of Primers in LAMP Detection of Chicken Infectious Anemia and Chicken Parvovirus

[0032] 1. Preparation of DNA Standards

[0033] Use PCR to amplify the target fragments of CAIV VP1 gene and ChPV NS1 gene respectively, recover and purify the product and connect it to pGM-T vector, and select positive clone bacteria by PCR, and extract the plasmid of the positive clone bacteria with a kit. The DNA concentration was measured by a nucleic acid detector, and the concentration was calculated as the copy number according to the relative molecular mass and Avogadro constant. Store at -70°C for later use.

[0034] 2. Optimizing the LAMP reaction system, reaction conditions and kit construction

[0035] The reaction system is 20 μL: 2 μL DNA template (including 1 μL each of CIAV and ChPV DNA template), 10× buffer 2.5 μL, MgSO4 (100 mM) 1.5 μL, dNTP Mix (10 mM) 3.5 μL, Bst 2.0DNA Polymerase (8000U / mL) 1 μL, 40 pmol each inner primer (CIAV-FIP, CIAV-BIP,...

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Abstract

The invention discloses a double LAMP detection primer group for chicken infectious anemia and chicken parvovirus. Primers have base sequences of sequence table SEQ.ID.No.1 to SEQ.ID.No.6. Based on the primers, by optimizing the reaction system and conditions, inventors also develop a corresponding kit and establish a corresponding dual LAMP method. The method can be completed only by performing aconstant-temperature reaction in a water bath at 63 DEG C for 90 min, the method can specifically detect the chicken infectious anemia and the chicken parvovirus, the detection sensitivity is very high, and the method can detect 10 mixed template copies / [mu]L of a chicken infectious anemia and chicken parvovirus DNA mixed sample. Tests confirm that the method provided by the invention has more specific and sensitive characteristics than a conventional detection method, and positive results are detected by a color of fluorophore, so that false positive results are effectively suppressed, and aerosol pollution is reduced; and the method has better application prospects.

Description

technical field [0001] The invention belongs to the technical field of virus detection, in particular to a double LAMP detection kit for chicken infectious anemia and chicken parvovirus. Background technique [0002] Chicken infectious anemia and chicken parenchyma are common in chicken flocks, and both are viral diseases that harm chicks. Chicken infectious anemia virus (Chichen infectious anemia virus, CIAV) can cause chicken bone marrow yellowing, lymphoid tissue atrophy, and then lead to hematopoietic function and immunosuppressive disorders in chickens. Chicken parvovirus (ChPV) mainly causes acute or chronic enteric diseases, malnutrition syndromes, stunting and stunting syndromes characterized by diarrhea, depression, thermoregulation disorders, growth retardation, and increased feed consumption. In recent years, countries and regions such as North America, Hungary, Brazil and South Korea have successively reported outbreaks of the disease. [0003] Traditional dete...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/6844C12N15/11
CPCC12Q1/701C12Q1/6844C12Q2531/119Y02A50/30
Inventor 谢芝勋方庆张艳芳张民秀罗思思谢丽基范晴曾婷婷黄娇玲谢志勤邓显文刘加波
Owner GUANGXI VETERINARY RES INST