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Recombined 3-type parainfluenza viruse nuclelc shell protein and process for preparing antibody thereof

A technology of nucleocapsid protein and parainfluenza, applied in the field of genetic engineering, can solve the problem of ineffective control of antibiotic abuse, affecting early diagnosis, early prevention and early treatment of parainfluenza virus infection, and limiting epidemiological investigation and research of human parainfluenza virus infection And other issues

Inactive Publication Date: 2006-06-28
首都儿科研究所
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This situation seriously affects the early diagnosis, early prevention and early treatment of parainfluenza virus infection in clinical practice, and cannot effectively control the abuse of antibiotics, which limits the epidemiological investigation and research of human parainfluenza virus infection

Method used

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  • Recombined 3-type parainfluenza viruse nuclelc shell protein and process for preparing antibody thereof
  • Recombined 3-type parainfluenza viruse nuclelc shell protein and process for preparing antibody thereof
  • Recombined 3-type parainfluenza viruse nuclelc shell protein and process for preparing antibody thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0059] Example 1 Obtaining of M1 antigenic fragment gene

[0060] 1. Design and synthesis of primers

[0061] According to the analysis of the antigenicity of NP protein by Antheprot software, the results are as follows figure 1 As shown, at the N-terminus of the parainfluenza virus NP protein, the 145-158, 181-247, 341-385 amino acid residue fragments are high antigenic regions, and the nucleotide sequence in the 436-1176bp interval is selected as the research object, and Primer Premier The software designed and synthesized the following primers containing BamH I and Xho I restriction endonuclease sites:

[0062] Upstream primer, 5' cgg gat cca gaa ggg cag aaa ca 3'

[0063] Downstream primer, 5' tcc gct cga ggc ttt ctt tgg ctt c 3'

[0064] 2. RT-PCR reaction to obtain the target gene fragment

[0065] Use Trizol to extract viral RNA from Vero cells infected with the standard strain of parainfluenza virus type 3, add 1 μL of Oligo dT with a concentration of 40 μg / μL to 9...

Embodiment 2

[0068] Example 2 Construction of recombinant pGEX-5X-3 / NP3-2 prokaryotic expression plasmid

Embodiment 2-1

[0069] Example 2-1 Construction of recombinant pGEX-5X-3 / NP3-2 prokaryotic expression plasmid

[0070] 1. Put 15 μL NP3-2 target gene fragment into a reaction system of 2 μL Xho I, 2 μL Buf-2, and 37°C for 12 hours, then add 2 μL BamH I, and continue digestion at 37°C for 2 hours;

[0071] 2. Put 6 μL pGEX-5X-3 into 2 μL BamH I, 2 μL Xho I, 4 μL Buf-3, 26 μL dHO 2 Carry out enzyme digestion in the reaction system at 0, 37°C, and let stand to react for 2 hours;

[0072] 3. Take 5 μL digested NP3-2 target gene fragment and 3 μL digested pGEX-5X-3 in 1 μL T4 DNA ligase, 1 μL 10×buffer, 13°C reaction system for 12 hours for ligation reaction to obtain the recombinant The pGEX-5X-3 / NP3-2 prokaryotic expression plasmid; extract the recombinant plasmid from the ampicillin-resistant transformed strain, cut the recombinant plasmid with BamH I and Xho I restriction endonucleases, and use 1% agarose gel Electrophoretic identification. The result is as figure 2 As shown in the figure...

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Abstract

The invention relates to bioengineering product manufacturing and application technique field, concretely relates to recombination 3 type parainfluenza virus core capsid protein, coding the protein gene and its manufacturing method and the application field of manufacturing antibody. The invention adopts molecular biology method to process in vitro recombination for antigen gene segment and manufacture the corresponding antigen protein to manufacture the corresponding antibody. It solves the defects in current technique that there is no simple and effective detecting system for clinic respiratory infected ill baby to do parainfluenza virus sieving-detecting and the import detecting kit is very expensive.

Description

technical field [0001] The invention relates to the technical field of genetic engineering, in particular to a recombinant type 3 parainfluenza virus (parainfluenza virus) nucleocapsid protein (Nuclear capsidprotein, NP) protein and its antibody, and their preparation method technical field. Background technique [0002] Parainfluenza virus is an important pathogen of respiratory tract infection in humans, especially children. It can cause pharyngitis, laryngitis, tracheitis and bronchitis, and severe cases can cause death due to throat obstruction. Parainfluenza virus is an RNA virus, and traditional virus isolation and culture are difficult and time-consuming; the timing of virus antibody diagnosis is not easy to grasp; polymerase chain reaction is sensitive, but it is not conducive to popularization in primary hospitals; Antigen diagnosis of influenza virus infection requires virus antibodies with strong specificity and high sensitivity. At present, in China, there is no...

Claims

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Application Information

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IPC IPC(8): C12N15/45C07K14/115C07K16/08C12N1/21C12N15/09C12N15/70
Inventor 辛若雷徐婧瑶张勤钱渊王芳张霆
Owner 首都儿科研究所
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