Mycoplasma ovipneumoniae multi-epitope fusion antigen MO-meAg5 and preparing method and application thereof

A Mycoplasma pneumoniae, mo-meag5 technology, applied in biochemical equipment and methods, bacterial antigen components, chemical instruments and methods, etc., can solve the problems that restrict the development of new vaccines for Mycoplasma ovis pneumonia and the complex antigen structure

Inactive Publication Date: 2016-08-17
SHIHEZI UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Due to the complex antigen structure of MO, no antigen with good immune protection has been screened and ident...

Method used

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  • Mycoplasma ovipneumoniae multi-epitope fusion antigen MO-meAg5 and preparing method and application thereof
  • Mycoplasma ovipneumoniae multi-epitope fusion antigen MO-meAg5 and preparing method and application thereof
  • Mycoplasma ovipneumoniae multi-epitope fusion antigen MO-meAg5 and preparing method and application thereof

Examples

Experimental program
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Embodiment 1

[0070] 1. Extraction of MO genome

[0071] A purified strain of pure bacteria was transferred to 30-50ml containing 15-20% heat-inactivated premium horse serum (Biotopped, China) (v / v), 0.004-0.06% phenol red (Sigma, USA) (w / v), 25~30μg / ml ampicillin (OMEGA, USA) in the brain heart infusion broth medium (BD, USA), at 36~38℃, the concentration is 5% CO 2 Cultivate in the incubator for 5-8 days, when the growth titer reaches 10 9 CCU / ml, centrifuge at 12000r / min for 15min, collect the bacteria, discard the supernatant, and then use the Mycoplasma gDNA Mini Kit (BIOMIGA, USA) extraction kit to extract the total DNA of Mycoplasma ovis pneumoniae.

[0072]2. Construction of MO genome expression library

[0073] Use Sau3A I enzyme (TaKaRa, Japan) to digest the total DNA of MO, and master the optimal amount and time of enzyme digestion through the enzyme digestion pre-experiment, so that the genome fragments digested by the enzyme are concentrated in 0.5 ~ 3kb, and then according ...

Embodiment 2

[0100] 1. Extraction of MO genome

[0101] Transfer the purified strain to 30-50ml culture medium, and incubate at 36-38°C with a concentration of 4-6% CO 2 Cultivate in the incubator for 5-9 days, when the growth titer reaches 10 9 CCU / ml and above, centrifuge at 10000-16000r / min for 10-20min, collect the bacteria, discard the supernatant, and then use the Mycoplasma gDNA Mini Kit (BIOMIGA, USA) extraction kit to extract the total DNA of Mycoplasma ovis pneumoniae.

[0102] The medium is Brain Heart Infusion Broth Medium (BD, USA).

[0103] The preferred composition of the culture medium is as follows: containing

[0104] Heat-inactivated premium horse serum (Biotopped, China) 15-20% (v / v),

[0105] Phenol red (Sigma, USA) 0.004~0.06% (w / v),

[0106] Ampicillin (OMEGA, USA) 25-30 μg / ml.

[0107] 2. Construction of MO genome expression library

[0108] Use Sau3A I enzyme (TaKaRa, Japan) to digest the total MO DNA, so that the digested genome fragments are concentrated in...

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Abstract

The invention discloses a mycoplasma ovipneumoniae multi-epitope fusion antigen MO-meAg5 and a preparing method and application thereof. Antigen protein high in immunogenicity namely the mycoplasma ovipneumoniae multi-epitope fusion antigen MO-meAg5 can be obtained by extracting an MO genome, constructing and screening an expression library, constructing the MO multi-epitope fusion antigen and expressing the multi-epitope fusion antigen in escherichia coli, and a foundation is laid for further researching and developing a safe and efficient mycoplasma ovipneumoniae multi-epitope protein vaccine.

Description

technical field [0001] The invention relates to an ovine mycoplasma pneumoniae multi-epitope fusion antigen MO-meAg5 and a preparation method and application thereof, belonging to the field of biotechnology. Background technique [0002] In recent years, there have been reports of the occurrence and prevalence of Mycoplasma ovis pneumonia in Xinjiang, Gansu, Inner Mongolia, Qinghai, Shaanxi, Ningxia, Liaoning, Jiangsu, Sichuan, Yunnan and other provinces with developed sheep farming industry. With the healthy development of the sheep raising industry, the harm is increasing day by day, causing huge economic losses to the local sheep raising industry. [0003] Mycoplasma ovipneumoniae (Mycoplasma ovipneumoniae, MO) belongs to Mollusca class, Mycoplasma order, Mycoplasma family, a kind of Mycoplasma genus. [0004] MO is a minimal prokaryotic microorganism that lacks cell walls, is highly pleomorphic, and can pass through bacteria filters. The MO genome is smaller than most ...

Claims

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Application Information

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IPC IPC(8): C07K14/30C12N15/70C40B50/06A61K39/02A61P31/04
CPCA61K39/0241C07K14/30C12N15/70C12N2800/101C12N2800/22C40B50/06
Inventor 乔军陈诚孟庆玲胡政香马玉田路路卢海亭曹旭东陈创夫
Owner SHIHEZI UNIVERSITY
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