Optimized DNA sequence, recombinant plasmid, strain, recombinant protein, mycoplasma gallisepticum antibody colloidal gold detection test paper and detection card

A technology of DNA sequence and Mycoplasma gallisepticum, which is applied in the directions of measuring devices, instruments, scientific instruments, etc., can solve problems such as colloidal gold test strips that have not disclosed Mycoplasma gallisepticum antibody, and achieve the effect of improving the detection effect.

Inactive Publication Date: 2019-07-26
HUAZHONG AGRI UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0008] In the process of realizing the present invention, the inventors have found that there are at least the following problems in the prior art: no colloidal gold test paper for antibodies against Mycoplasma gallisepticum has been disclosed, which can be used to quickly and simply detect the immunity level of birds to Mycoplasma gallisepticum

Method used

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  • Optimized DNA sequence, recombinant plasmid, strain, recombinant protein, mycoplasma gallisepticum antibody colloidal gold detection test paper and detection card
  • Optimized DNA sequence, recombinant plasmid, strain, recombinant protein, mycoplasma gallisepticum antibody colloidal gold detection test paper and detection card
  • Optimized DNA sequence, recombinant plasmid, strain, recombinant protein, mycoplasma gallisepticum antibody colloidal gold detection test paper and detection card

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Effect test

Embodiment 1

[0082] 1. Preparation of MG-VlhA1.2 recombinant protein

[0083] 1.1 Acquisition and optimization of Mycoplasma gallisepticum VlhA1.2 gene and construction of recombinant plasmid pGEX-KG-VlhA1.2

[0084] The method for obtaining and optimizing the sequence of the Mycoplasma gallisepticum VlhA1.2 gene involved in the present invention and the method for constructing the recombinant plasmid pGEX-KG-VlhA1.2 Expression [J], Chinese Journal of Veterinary Medicine, 2007.

[0085] The specific implementation method is as follows: obtain the VlhA1.2 gene sequence according to the pMGA1.2 sequence included in GenBank (accession number: AF275312). Since the UGA codon in Mycoplasma encodes tryptophan, and UGA is a stop codon in Escherichia coli, in order to achieve high-efficiency expression of the target gene in Escherichia coli, TGA was optimized to TGG; The gene was optimized according to the preference of Escherichia coli, and an EcoR I (GAATTC) restriction site was added to the 5'...

Embodiment 2

[0139] Evaluation of the stability of the colloidal gold detection test paper for mycoplasma fowl septicemia antibody of the present invention

[0140] The vacuum-packaged test papers of the present invention placed at 4°C and 37°C respectively were taken out on the 7th, 16th, 35th, 54th, 70th, and 90th day, and tested after the standard positive serum of Mycoplasma gallisepticum was diluted with pure water. . The results are shown in Table 2. The test paper of the present invention has a longer shelf life at 4° C., and the shelf life is at least 90 days.

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Abstract

The invention provides an optimized DNA sequence, a recombinant plasmid, a strain, a recombinant protein, mycoplasma gallisepticum antibody colloidal gold detection test paper containing the recombinant protein and a detection card based on mycoplasma gallisepticum VlhA1.2 protein, wherein the test paper can be used for quickly detecting diagnosis of poultry after mycoplasma gallisepticum infection and antibody level detection after immunization. The optimized DNA sequence is shown as SEQ ID NO.1. The optimized DNA sequence is combined with a pGEX-KG vector to obtain the recombinant plasmid pGEX-KG-VlhA1.2, and transforming and expressing is carried out to obtain the MG-VlhA1.2 recombinant protein. The test paper comprises a gold-labeled pad and a nitrocellulose membrane which are sequentially connected, wherein the gold-labeled pad is coated with a colloidal gold label of a monoclonal antibody mouse anti-IgY Fc CH3-CH4; and one end of the nitrocellulose membrane, which is close to thegold-labeled pad, is a detection line coated with MG-VlhA1.2 recombinant protein.

Description

technical field [0001] The invention relates to the technical field of animal immunity application, in particular to an optimized DNA sequence, recombinant plasmid, bacterial strain, recombinant protein, Mycoplasma gallisepticum antibody colloidal gold detection test paper and detection card. Background technique [0002] Mycoplasma gallisepticum is an infectious disease caused by Mycoplasma gallisepticum (MG), also known as Chronic Respiratory Disease (CRD), which is prevalent in all poultry-raising countries in the world, and the infection rate in my country is also very high. Some areas reach more than 75%. MG has a wide range of hosts: chickens, quails, pheasants, ducks, pigeons, geese, peacocks and cranes can be infected with MG. The disease can be transmitted vertically and horizontally, which is also an important reason for the high infection rate, wide distribution and difficulty in eradication of Mycoplasma gallisepticum. No matter in developed countries or in deve...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N33/569G01N33/532G01N33/543
CPCG01N33/532G01N33/54306G01N33/54313G01N33/56983
Inventor 李自力邵雨胡思顺毕丁仁周祖涛刘梅石德时许青荣
Owner HUAZHONG AGRI UNIV
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