Tubercle bacillus fusion protein HspX-Mtb8.4 and preparation method and application thereof

A technology of hspx-mtb8.4 and fusion protein, which is applied in the direction of chemical instruments and methods, bacteria, hybrid peptides, etc., can solve unconsidered and affected problems, and achieve the effect of strong cellular immune activity

Inactive Publication Date: 2011-10-12
LANZHOU UNIVERSITY
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  • Claims
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Problems solved by technology

However, in previous studies, fusion proteins with various tags (such as His.Tag, GST.Tag, S.Tag, etc.) were often constructed for the convenience of purification methods in the later stage, but the fusion protein was not considered. Whether the label will affect the animal experiment and further

Method used

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  • Tubercle bacillus fusion protein HspX-Mtb8.4 and preparation method and application thereof
  • Tubercle bacillus fusion protein HspX-Mtb8.4 and preparation method and application thereof
  • Tubercle bacillus fusion protein HspX-Mtb8.4 and preparation method and application thereof

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Experimental program
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Embodiment 1

[0047] 1. Construction of recombinant plasmids

[0048] The purified PCR products Mtb8.4 and Hspx and plasmid PET30a were digested with restriction endonucleases Nde I and Sac I respectively, and the digested Mtb8.4T and Hspx gene fragments and plasmid PET30a DNA fragments were passed through T4 DNA ligase After connecting overnight at 16°C, transform into Escherichia coli strain DH5α, spread on LB plates containing 50 mg / mL kanamycin, culture overnight at 37°C, pick positive clones, extract plasmids, and perform PCR and sequencing Identification. The recombinant plasmids PET30a-Mtb8.4 and PET30a-Hspx extracted and sequenced correctly were digested with restriction endonucleases Sac I and HindIII respectively, and Hspx and Mtb8.4 were digested with Sac I and HindIII respectively. After gel recovery and purification, the target gene fragments were ligated with the recovered products of PET30a-Mtb8.4 and PET30a-Hspx vectors under the action of T4 DNA ligase at 16°C overnight to...

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Abstract

The invention discloses a tubercle bacillus fusion protein HspX-Mtb8.4 which is a protein shown as a sequence 2 in a sequence table. The invention has the advantages that: a mycobacterium tuberculosis Mtb8.4 is a low molecular treatment protein antigen purified and separated from tubercle bacillus culture filtrate, has high cellular immune activity, and can be used for inducing a CD4+TH1 cellular immune reaction and the generation of a specific CTL (Cell Toxicity Lymphocyte) and secreting high-level IFN (Interferon)-gamma. HspX is a heat shock protein with the relative molecular mass of 16,000 and is a major protein produced by mycobacterium tuberculosis at a stationary growth phase or in a low-oxygen state. Specific humoral immunity and cellular immune response specific to HspX in a patient suffering from tuberculosis and a latent-phase infected person indicate that HspX is an important target antigen of an organism immune reaction at a latent infective phase of mycobacterium tuberculosis.

Description

technical field [0001] The invention relates to a Mycobacterium tuberculosis fusion protein HspX-Mtb8.4 and its preparation method, as well as its application in the preparation of candidate vaccines against tuberculosis. Background technique [0002] Tuberculosis is one of the serious diseases that endanger human health for a long time. Nearly 1 / 3 of the people in the world are still infected with Mycobacterium tuberculosis. About 5% of tuberculosis infected patients will develop tuberculosis patients within 2-5 years; the rest may form latent tuberculosis infections. my country is one of the countries with a high burden of tuberculosis, and about 130,000 people die of tuberculosis every year. At present, BCG vaccination is an important measure to effectively prevent tuberculosis. However, different studies have shown that the protective effect of BCG in different populations is not stable, especially the protective efficiency of adult pulmonary tuberculosis ranges from ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K19/00C12N15/62C12N15/63C12N1/21A61K39/04A61P31/06
Inventor 祝秉东刘万波谭继英胡丽娜王秉翔吴玉敏辛奇朱丽林孝发
Owner LANZHOU UNIVERSITY
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