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Soluble expression method of recombinant sheeppox virus fusion protein and biological material used therefor

A sheep pox virus, fusion protein technology, applied in the direction of biological material analysis, virus, virus peptides, etc., can solve the problem of poor sensitivity, and achieve the effect of high specificity, simple operation and good sensitivity

Active Publication Date: 2020-05-08
CHINA ANIMAL DISEASE CONTROL CENT
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Dr. Ma Weimin of Lanzhou Veterinary Research Institute established the iELISA method using the main antigenic protein P32 of sheeppox virus in 2006. It has good specificity and no cross-reaction with sheep's mouth sore and vaccinia infected sera, but its sensitivity is poor

Method used

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  • Soluble expression method of recombinant sheeppox virus fusion protein and biological material used therefor
  • Soluble expression method of recombinant sheeppox virus fusion protein and biological material used therefor
  • Soluble expression method of recombinant sheeppox virus fusion protein and biological material used therefor

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0078] Example 1. Soluble expression of recombinant sheeppox virus fusion protein

[0079] 1. Construction of recombinant expression vector

[0080] 1.1 Construction of recombinant sheeppox virus fusion protein gene recombinant expression vector by fusion of three sheeppox virus protein fragments

[0081] This application has designed three recombinant sheeppox virus fusion protein genes, which are composed of three sheeppox virus protein fragments, which are His-L1-P32-A33-Y-His gene (CDS position 4-1917), the His-L1-P32-A33-W-His gene shown in SEQ ID No. 3 (position 4-1917 in CDS) (as the His-L1-P32-A33-Y-His gene control) and the His-obL1-P32-A33-Y-His gene shown in SEQ ID No. 4 (CDS is position 4-2436) (as a control for the His-L1-P32-A33-Y-His gene) . Both His-L1-P32-A33-Y-His gene and His-L1-P32-A33-W-His gene encode the protein His-L1-P32-A33-Y-His shown in SEQ ID No. Invented recombinant sheeppox virus fusion protein). The His-obL1-P32-A33-Y-His gene encodes the p...

Embodiment 2

[0115] Example 2. Detection of Sheeppox Virus Antibody by Indirect ELISA Method Using His-L1-P32-A33-Y-His Protein as Coating Antigen

[0116] The relevant solutions in this embodiment are as follows:

[0117] Preparation of PBS buffer with a concentration of 0.01M and a pH of 7.4: 8.5g NaCl, 0.2g KCl, 2.9gNa 2 HPO 4 ·12H 2 O, 0.59g NaH 2 PO 4 ·2H 2 O, 1L deionized water.

[0118] Coating buffer: 0.05mol / L sodium carbonate-sodium bicarbonate buffer (pH9.6), the solvent is water, the solute and its concentration are as follows: Na 2 CO 3 1.59g / L and NaHCO 3 2.93g / L.

[0119] Washing solution: 0.5% Tween washing solution. The 0.5% Tween washing solution is prepared according to the following method: Tween 20 and sodium azide are added to the PBS buffer with a concentration of 0.01M and a pH value of 7.4 until the content of sodium azide is 5g / L, Tween 20 The content of 5mL / L, to obtain 0.5% Tween washing solution.

[0120] Blocking solution: 1% BSA blocking solutio...

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Abstract

The invention discloses a soluble expression method for a recombinant sheep pox virus fusion protein and a biological material used in the soluble expression method. The biological material is used for preparing a protein a), b) or c), wherein a) is a protein of which the amino acid sequence is an 8th-629th amino acid residue as shown in SEQ ID No.2; b) is a protein of which the amino acid sequence is as shown in SEQ ID No. 2; and c) is a soluble protein which is obtained by substituting and / or deleting and / or adding one or more amino acid residues on an amino acid sequence as shown in SEQ IDNo.2 and has the activity same as that of a). The method for indirectly detecting a sheep pox virus antibody by an ELISA, established by taking the recombinant sheep pox virus fusion protein preparedfrom the biological material as an envelope antigen, not only can be used for diagnosing sheep pox, but also can be used for evaluating a vaccine immunity effect; in addition, the method can be used for rapidly and accurately detecting the sheep pox so as to play an active role in better controlling the propagation of the sheep pox in China.

Description

technical field [0001] The invention relates to a soluble expression method of recombinant sheeppox virus fusion protein in the field of biotechnology and biological materials used therefor. Background technique [0002] Sheeppox, also known as sheep "smallpox", is an acute, febrile, and highly contagious infectious disease caused by Capripoxvirus (CaPV). It is listed as a notifiable infectious disease by the World Organization for Animal Health (OIE), which shows its importance. Due to the acute onset and high incidence of the disease, the quality of wool and cashmere and the output of meat products will be greatly affected after the onset of the disease, which has caused huge economic losses to my country's sheep farming industry. At present, the standard method for detecting anti-sheeppox virus antibodies is the sheeppox serum neutralization test, which is an effective method for detecting sheeppox virus antibodies, but there are also many shortcomings. The main disadva...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C07K19/00C12N15/62C12N15/70G01N33/577G01N33/569
CPCC07K14/005C07K2319/00C12N15/70C12N2710/24222C12N2800/22G01N33/56983G01N33/577G01N2469/20
Inventor 孙雨宋晓晖杨林王传彬马英毕一鸣杨天意张晨
Owner CHINA ANIMAL DISEASE CONTROL CENT
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