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Package for identifying canine parvovirus and kit

A parvovirus and dog identification technology, applied in the biological field, can solve problems such as inconvenient on-site detection, and achieve the effects of convenient virus detection, good specificity and simple operation

Inactive Publication Date: 2020-02-21
北京美莱博医学科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, conventional LAMP amplification still requires special detection equipment, which is not convenient for on-site detection requirements

Method used

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  • Package for identifying canine parvovirus and kit
  • Package for identifying canine parvovirus and kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Embodiment 1, the preparation of the complete set of reagents of identifying canine parvovirus

[0037] The set of reagents for identifying canine parvovirus provided in this example is composed of six single-stranded DNAs named CPV-F3, CPV-B3, CPV-FIP, CPV-BIP, CPV-FLP and CPV-BLP, each single-stranded DNA The sequence is as follows:

[0038] CPV-F3: 5'-TGGGATAAAGAATTTGATACTGA-3' (sequence 1 in the sequence listing);

[0039] CPV-B3: 5'-TGAGAGGCTCTTAGTTTAGCTTT-3' (sequence 2 in the sequence listing);

[0040] CPV-FIP: 5'-CGCAACTTTTACAAATAATTGACCATTAAAACCAAGACTTCATGTAAATGC-3' (sequence 3 in the sequence listing);

[0041] CPV-BIP: 5'-AACAAATGAATATGATCCTGATGCAACCTTTCCACCAAAAATCTGAGT-3' (sequence 4 in the sequence listing);

[0042] CPV-FLP: 5'-GGACAATTATTTTGACAAACAAATGG-3' (sequence 5 in the sequence listing);

[0043] CPV-BLP: 5'-TCTGCTAATATGTCAAGAATTGTAAC-3' (SEQ ID NO: 6 in the Sequence Listing).

[0044] In this set of reagents, CPV-F3, CPV-B3, CPV-FIP, CPV-BIP, C...

Embodiment 2

[0045] Embodiment 2, establishment of canine parvovirus identification method

[0046] The 25 μL reaction system for the identification of canine parvovirus contains: 2 μl of canine parvovirus genomic DNA, 10 mM (NH 4 ) 2 SO4 (Amresco, USA), 50mM KCl (Amresco, USA), 8mM MgSO 4 (Beijing Meilaibo Medical Technology Co., Ltd.), 50uM neutral red (Sigma, USA), 0.1% 20 (U.S. Amresco company), 0.5M betaine (U.S. Sigma company), 1.4mM dNTPs (Beijing Meilaibo Medical Technology Co., Ltd.), CPV-F3, CPV-B3, CPV-FIP, CPV-BIP of embodiment 1 , CPV-FLP and CPV-BLP, Bst2.0 DNA polymerase (NEB Company) 0.4U / μl, and 25 μl of sterile water. The concentrations of CPV-F3, CPV-B3, CPV-FIP, CPV-BIP, CPV-FLP and CPV-BLP in this system were 0.2μM, 0.2μM, 1.6μM, 1.6μM, 0.4μM and 0.4μM, respectively.

[0047] Incubate the above-mentioned canine parvovirus-identified reaction systems at 65° C. for 40 min to carry out LAMP. According to the color change of the system, determine whether the sample t...

Embodiment 3

[0048] Example 3, Identification of the specificity of the set of reagents for canine parvovirus

[0049] Canine parvovirus DNA, Staphylococcus aureus (ATCC 6538) DNA, Shigella dysenteriae (Shigella dysenteriae ATCC 51252) DNA, Enterobacter sakazakii (ATCC 51329) DNA, and Les monocytogenes were used respectively at a concentration of 80ng / μl. Special bacteria (ATCC19114) DNA and Escherichia coli (ATCC 25922) DNA (provided by Zhejiang Quality Inspection Research Institute) were used as templates for LAMP reaction, and LAMP was carried out according to the reaction system and reaction conditions in Example 2, and real-time monitoring was carried out. Each strain was replicated in triplicate. Set up a negative control by replacing the template with sterile water.

[0050] Among them, canine parvovirus is recorded in the article "Zhao Guoxing et al., Establishment of indirect ELISA detection method for canine parvovirus antibody, Chinese Journal of Veterinary Medicine, September ...

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Abstract

The invention discloses a package for identifying canine parvovirus and a kit. The package for identifying the canine parvovirus, disclosed by the invention, is prepared from six single-stranded DNAs,i.e., CPV-F3, CPV-B3, CPV-FIP, CPV-BIP, CPV-FLP and CPV-BLP; and sequences of the CPV-F3, CPV-B3, CPV-FIP, CPV-BIP, CPV-FLP and CPV-BLP are separately sequences 1-6 in a sequence listing. Proven by experiments, the identification on the canine parvovirus by using the package for identifying the canine parvovirus, disclosed by the invention, is convenient and rapid and is good in specificity and high in sensitivity.

Description

technical field [0001] The invention relates to a set of reagents and a kit for identifying canine parvovirus in the field of biotechnology. Background technique [0002] Canine parvovirus (CPV) belongs to the family Parvoviridae and was first isolated in 1978 from the feces of dogs suffering from enteritis; CPV infection can cause canine parvovirus enteritis, which is one of the severe infectious diseases of dogs worldwide. Its characteristics of short course of disease, strong infectivity and high mortality are very harmful to dogs, and fast and accurate diagnostic methods are needed for timely treatment. [0003] The rapid detection of CPV mainly includes immunological detection based on serum protein and genetic detection for viral DNA. Genetic testing includes routine PCR amplification and fluorescent quantitative PCR testing. [0004] LAMP amplification is a constant temperature amplification method invented by Notomi et al. in Japan. It is characterized in that it r...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/6844C12N15/11
CPCC12Q1/701C12Q1/6844
Inventor 李洪波廖翔
Owner 北京美莱博医学科技有限公司