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Adenovirus high expression vector for improving expression of hantavirus fusion protein G1S0.7

A technology of fusion protein and hantavirus, applied in the direction of using vectors to introduce foreign genetic material, application, biochemical equipment and methods, etc., can solve the problem that genes cannot be expressed satisfactorily

Inactive Publication Date: 2010-08-25
FOURTH MILITARY MEDICAL UNIVERSITY
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Problems solved by technology

Although the widely used CMV promoter is a relatively effective expression system promoter, some genes usually cannot be expressed satisfactorily in practical applications.

Method used

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  • Adenovirus high expression vector for improving expression of hantavirus fusion protein G1S0.7
  • Adenovirus high expression vector for improving expression of hantavirus fusion protein G1S0.7
  • Adenovirus high expression vector for improving expression of hantavirus fusion protein G1S0.7

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Embodiment Construction

[0020] The adenovirus expression system used in the present invention was purchased from Adeno-X of CLONTECH company TM system (Product No. K1650-1), the transfer vector is pShuttle, which contains CMV promoter.

[0021] The Hantaan virus used in the present invention is Hantaan virus 76-118 strain. Its genome consists of three gene segments, L, M and S. The L segment encodes RNA-dependent RNA polymerase; the M segment encodes envelope glycoproteins (GP) G1 and G2; the S segment encodes nucleocapsid protein (NP). The fusion gene involved in the present invention is obtained by PCR amplification of the G1 part of the M segment and the 0.7 part of the S segment respectively, and is fused by gene fusion technology, and Xba I is designed at the 5' end and Not at the 3' end. I restriction site, cloned into the corresponding restriction site of the adenovirus transfer vector pShuttle to obtain a recombinant plasmid, named G1S0.7-pShuttle.

[0022] The replaced CAG promoter / enhanc...

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Abstract

The invention relates to reconstruction of an adenovirus transfer vector, in particular to reconstruction of a transfer vector of mosaic gene G1S0.7 of hantavirus (HV) serving as hemorrhagic fever with renal syndrome (HFRS) pathogen for improving the expression of hantavirus fusion protein G1S0.7. The adenovirus transfer vector G1S0.7-pShuttle containing the mosaic gene G1S0.7 is reconstructed by using the gene recombination technology. The reconstruction comprises the following steps: replacing a CAG promoter / enhancer for a CMV promoter; or inserting a WPRE transcriptional control element at the 3' end of the promoter; or replacing the promoter and inserting a WPRE control element. The vector expresses the fusion proteins G1S0.7 respectively, and compares the expression level of foreign protein in each recombinant adenovirus. The result shows that the vector can effectively improve the expression of the fusion protein G1S0.7 compared with the primary transfer vector pShuttle, and has most obvious effect of replacing a promoter group.

Description

technical field [0001] The invention belongs to the field of biotechnology, and relates to related fields such as molecular biology, immunology and immune application. The invention relates to the reconstruction of the adenovirus recombinant transfer vector and its expression to the chimeric gene G1S0.7 of Hantavirus (HV), the pathogen of hemorrhagic fever with renal syndrome (HFRS). Background technique [0002] Research status of hemorrhagic fever with renal syndrome and its genetic engineering vaccine: [0003] Hemorrhagic fever with renal syndrome (HFRS) is caused by Hantavirus (HV) and is an acute viral infectious disease transmitted by rodents. Clinically, it is characterized by fever, bleeding and acute renal failure. Damage is the main feature. China is the country with the most severe HFRS epidemic in the world. It has the characteristics of a wide range of epidemics, a large number of patients, and a high mortality rate. So far, there is still a lack of specific ...

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Application Information

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IPC IPC(8): C12N15/861C12N15/67
Inventor 徐志凯张芳琳白文涛李璞媛吴兴安李凯胡刚
Owner FOURTH MILITARY MEDICAL UNIVERSITY
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