A vector for expressing poliovirus-like particle protein and preparation method of poliovirus-like particle

A poliomyelitis and granule protein technology, applied in the fields of biotechnology and biomedicine, can solve the problems of large IRES structure, limited carrier capacity, reduced transfection efficiency, etc. Effect

Active Publication Date: 2017-07-04
SOUTH CHINA UNITED VACCINE INST
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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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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  • A vector for expressing poliovirus-like particle protein and preparation method of poliovirus-like particle
  • A vector for expressing poliovirus-like particle protein and preparation method of poliovirus-like particle
  • A vector for expressing poliovirus-like particle protein and preparation method of poliovirus-like particle

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preparation example Construction

[0054] A method for preparing a vector expressing poliovirus-like particle protein, comprising 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 i...

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

Disclosed are a vector expressing poliomyelitis virus-like granule proteins and a method for preparing poliomyelitis virus-like granules. The vector contains expression cassettes of the following structure: any one of three genes of poliomyelitis virus structural proteins i.e. VP0, VP1, VP3, is located in the downstream of promoter 1, and the other two are connected to the downstream of promoter 2 by a 2A sequence. The promoting expressions directions of the two promoters are opposite. The method for preparing poliomyelitis virus-like granules is that the corresponding host cells are transfected with the vector. After cultivation, virus-like granules and recombinant baculoviruses can be obtained. If what are obtained are the recombinant baculoviruses, then the host cells are infected with the viruses, and virus-like granules can be obtained.

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 Patents(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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