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Application of protein Mip in immune protection of coxiella burnetii

A protein and protective antigen technology, applied in the direction of blood/immune system cells, antibacterial drugs, vertebrate cells, etc., can solve the problems of low reproductive capacity of chicken embryos, high protection requirements, high cost, etc., to overcome preparation difficulties, The effect of simple operation and low cost

Inactive Publication Date: 2013-05-08
MICROBE EPIDEMIC DISEASE INST OF PLA MILITARY MEDICAL ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Both inactivated vaccines and chemically extracted subunit vaccines require a large number of chicken embryos to multiply Coxiella beinii for preparation, but the number of chicken embryos is low, and the protection requirements for extracting and purifying Coxiella beinii are high, the process is complicated, and the cost is expensive , so it is difficult to prepare and promote the use of large quantities

Method used

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  • Application of protein Mip in immune protection of coxiella burnetii
  • Application of protein Mip in immune protection of coxiella burnetii
  • Application of protein Mip in immune protection of coxiella burnetii

Examples

Experimental program
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Effect test

Embodiment 1

[0032] Embodiment 1, the preparation of antigenic protein Mip

[0033] The vector pET-32a(+) was purchased from Novagen, the product catalog number is 69015. Escherichia coli BL21 was purchased from Novagen, the product catalog number is 69450.

[0034] Preparation of the coding gene of protein Mip: using the genomic DNA of Coxiella burnetii as a template, carry out PCR amplification with primer pairs to obtain PCR amplification products; use restriction endonucleases BamHI and XhoI Enzyme digestion of the PCR amplification product to recover the target gene fragment; use restriction endonucleases BamHI and XhoI to digest the vector pET-32a(+) to recover the large vector fragment; connect the target gene fragment and the large vector fragment to obtain the recombinant vector; PCR amplification conditions: 94°C pre-denaturation for 5 minutes, 35 cycles at 95°C for 30 sec, 55°C for 30 sec, 72°C for 1.5 min, and 72°C extension for 7 min.

[0035] 5'ATA GGATCC ATGAAACGATTGATTT...

Embodiment 2

[0053] Embodiment 2, the application of antigenic protein Mip

[0054] 1. Preparation of protective antigen against Q fever

[0055] 1. Preparation of immature dendritic cells

[0056] BALB / c mice were purchased from the Experimental Animal Center of the Academy of Military Medical Sciences.

[0057] Isolate bone marrow cells from the femoral bone marrow of BALB / c mice: Take the femur of BALB / c mouse, cut off the femoral heads on both sides with ophthalmic scissors, insert the syringe needle filled with 1ml of PBS buffer into the bone cavity, and slowly push the syringe to rinse , Flush a single bone marrow cell suspension from the bone cavity.

[0058] Adjust the number of bone marrow cells to 1×10 with RPMI1640 full medium 6 / ml, and transferred to a six-well culture plate (9.6×6cm 2 ), cultured at a temperature of 37°C and a carbon dioxide volume concentration of 5%, on the 3rd and 5th days of culture, each well was replaced with 3ml of RPMI1640 full medium, and culture...

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Abstract

The invention discloses application of protein Mip in the immune protection of coxiella burnetii. The application means the application of protein shown as SEQ IDNO: 1 in the preparation of a Q fever vaccine. The application proves that coxiella burnetii recombinant membrane protein Mip can be used as antigenic protein, can stimulate bodies to produce immune response to attack anti-coxiella burnetii strains and is a protective antigen with Q fever vaccine functions. The protein Mip is prepared by a genetic engineering method, and the method is easy to operate, low in cost and high in yield. The protein Mip is used as the Q fever vaccine, so that the defect of difficulty in preparation of vaccines in the prior art is overcome.

Description

technical field [0001] The present invention relates to the application of protein Mip in immune protection against Coxiella bezieri. Background technique [0002] Coxiella burnetii is a small Gram-negative bacillus parasitic in obligate phagocytic cells. The bacterium has strong resistance to the external environment and has strong infectivity to humans and animals. It can enter the body through aerosol diffusion through the respiratory tract, causing infection in humans and animals and Q fever. [0003] Q fever is a zoonotic disease with worldwide distribution and is widely distributed in my country. The clinical manifestations of Q fever in humans are diverse. Acute Q fever manifests as fever, headache, muscle aches, often accompanied by pneumonia, hepatitis, etc.; chronic Q fever manifests as endocarditis, granulomatous hepatitis, osteomyelitis, etc. In recent years, Q fever meningitis, glomerulonephritis, and abortion in pregnant women have also been reported. Altho...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): A61K39/02C12N5/0784A61P31/04
Inventor 熊小路温博海王锡乐
Owner MICROBE EPIDEMIC DISEASE INST OF PLA MILITARY MEDICAL ACAD OF SCI
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