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Vector for expressing protein of poliovirus sample particles and method for preparing poliovirus sample particles

A polio and granular protein technology, applied in the fields of biotechnology and biomedicine, can solve the problems of large IRES structure, limited carrier capacity, and increased carrier burden, and achieve good immunogenicity, simple operation, and high expression.

Active Publication Date: 2015-04-01
SOUTH CHINA UNITED VACCINE INST
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

The system has the following defects: (1) IRES itself has a large structure (about 0.5kb), its application is often limited by the capacity of the vector, and it will increase the burden of the vector, resulting in a decrease in transfection efficiency
Therefore, in the construction of polycistronic vector system, IRES is not an ideal connecting element

Method used

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  • Vector for expressing protein of poliovirus sample particles and method for preparing poliovirus sample particles
  • Vector for expressing protein of poliovirus sample particles and method for preparing poliovirus sample particles
  • Vector for expressing protein of poliovirus sample particles and method for preparing poliovirus sample particles

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preparation example Construction

[0054] Preparation method of a vector expressing poliovirus-like particle protein , including the following steps:

[0055] 1) According to the P1 gene sequence of poliovirus, codon optimization is carried out according to the preference of the host cell, and the optimized P1 gene sequence is synthesized;

[0056] 2) According to the optimized P1 gene sequence, design primers to clone any one of the VP0, VP1, and VP3 genes into the backbone vector, located downstream of a promoter of the backbone vector, and then connect the other two genes through the 2A sequence, And the ligated fragment is cloned to the downstream of another promoter of the same backbone vector; the resulting recombinant vector is the vector for expressing the poliovirus-like particle protein.

[0057] Preferably, the above-mentioned host cells are Spodoptera frugiperda Spodoptera frugiperda cells Sf9, Saccharomyces cerevisiae or mammalian cells.

[0058] Preferably, the above-mentioned skeleton vector ...

Embodiment 1

[0089] Example 1 A vector expressing poliovirus-like particle protein

[0090] Place the VP1 sequence of poliovirus type I in the P of the pFastBac Dual plasmid 10 Under the promoter, one end of the 2A sequence gene of Picornaviridae (i.e., the encoded amino acid sequence is GSGATNFSLLKQAGDVEENPGP) is connected to the VP3 gene of poliovirus type I, and the other end is connected to the VP3 gene of poliovirus type I VPO gene, placed in the P of the pFastBac Dual plasmid polh Under the promoter, the shuttle vector pFBD-I VP1-VP3-2A-VPO is obtained, which is a vector for expressing poliotype I virus-like particle protein.

Embodiment 2

[0091] Example 2 A vector expressing poliovirus-like particle protein

[0092] Place the VP1 sequence of poliovirus type II in the P of the pFastBac Dual plasmid 10 Under the promoter, one end of the 2A sequence gene of Picornaviridae (i.e., the encoded amino acid sequence is GSGATNFSLLKQAGDVEENPGP) is connected to the VP3 gene of type II poliovirus, and the other end is connected to the VP3 gene of type II poliovirus VPO gene, placed in the P of the pFastBac Dual plasmid polh Under the promoter, the shuttle vector pFBD-II VP1-VP3-2A-VPO is obtained, which is a vector for expressing poliotype II virus-like particle protein.

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Abstract

The invention discloses a vector for expressing protein of poliovirus sample particles and a method for preparing the poliovirus sample particles. The vector contains expression cassettes with the following structures: an arbitrary gene in poliovirus structural proteins VP0, VP1 and VP3 genes is located in the downstream of a promoter 1, and the other two genes are connected by virtue of a 2A sequence and are located in the downstream of a promoter 2, and directions of promoting expressions of the two promoters are opposite. The method for preparing the poliovirus sample particles comprises the following steps: transfecting the carrier to a corresponding host cell, culturing to obtain virus particles or recombinant baculovirus, and infecting the virus to the host cell if the recombinant baculovirus is obtained so as to obtain the virus sample particles. The poliovirus sample particles can be used for respectively inducing high-titer neutralization titer in a body, and can be used as a vaccine for preventing and treating poliovirus infected related diseases.

Description

technical field [0001] The invention belongs to the fields of biotechnology and biomedicine, and relates to a carrier for expressing poliovirus-like granule protein and a preparation method for poliovirus-like granule. Background technique [0002] Poliomyelitis is an acute intestinal infectious disease mainly caused by poliovirus infection, which mainly affects limb paralysis. It mainly affects children under the age of five. There is no specific treatment for poliomyelitis, and it is widely spread worldwide and extremely harmful Because of the specific pathological changes and the damage of gray and white matter cells in the anterior horn of the spinal cord, especially in the gray matter area, it is called polio. The clinical feature is muscle paralysis, especially the flaccid paralysis of the body, which mostly occurs in children under 5 years old, especially infants, so it is also called polio (niafntelipaarylsis), but it does not only affect young children, but also oft...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/866C12N7/04A61K39/13A61P31/14
CPCA61K39/13C12N7/04C12N15/66C12N15/866Y02A50/30
Inventor 彭涛许煜华马书智王弋安鸿尹海滨
Owner SOUTH CHINA UNITED VACCINE INST
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