Rabbit hemorrhagic disease virus-like particles as well as preparation method and application thereof

A technology for rabbit hemorrhagic virus and rabbit hemorrhage, which is applied in the fields of microorganisms and veterinary biopharmaceuticals, can solve the problems of unsatisfactory expression level, unoptimistic application prospect, insolubility of VP60 and poor immune effect, etc., and achieves low cost and safety. High, immunogenic effect

Active Publication Date: 2014-02-05
CHANGCHUN SR BIOLOGICAL TECH
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0007] At present, there are still some problems in the research of RHDV genetically engineered vaccines: VP60 expressed by Escherichia coli has the disadvantages of insolubility and relatively poor immune effect, and its application prospects are not optimistic; Problem; the expression level of VP60 produced by transgenic plants is not ideal at present; therefore, it is imperative to develop a safe and efficient new vaccine for RHD

Method used

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  • Rabbit hemorrhagic disease virus-like particles as well as preparation method and application thereof
  • Rabbit hemorrhagic disease virus-like particles as well as preparation method and application thereof
  • Rabbit hemorrhagic disease virus-like particles as well as preparation method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0051] Example 1 Expression and Detection of Single Copy Rabbit Hemorrhagic Disease Virus VP60 Gene in Recombinant Baculovirus

[0052] 1. Experimental method

[0053] 1.1. Preparation of related solutions and medium

[0054] Solution I: 50mmol / L glucose, 25mmol / L Tris-HCl (pH8.0), 10mmol / L EDTA.

[0055] Solution II: 0.2mol / L NaOH, 1% SDS (preparation for current use).

[0056] Solution III: 100mL system, 80mL 5mol / L potassium acetate, 12mL glacial acetic acid, 8mL ddH2O.

[0057] TAE (50×): 242g Tris base, 57.1mL glacial acetic acid, 100mL0.5mol / L EDTA (pH8.0), dilute to 1000mL with sterile water.

[0058] TER solution: Pancreatic RNAse (RNAse A) was dissolved in 10mM Tris-HCl, 15mMNaCl, prepared as a 10mg / mL storage solution and stored at -20°C, and diluted with 1×TE buffer to a 20μg / mL working solution and stored at 4°C.

[0059] PPt buffer: isopropanol 22mL; 5mol / mL KAc1mL; ddH2O2mL.

[0060] 1×TE buffer solution: 10mmol / L Tris·Cl (pH8.0), 1mmol / L EDTA (pH8.0), steri...

Embodiment 2

[0152] Example 2 High-efficiency expression and application of double-copy VP60 optimized gene in baculovirus-insect cells

[0153] 2 Experimental methods

[0154] The preparation of relevant solution and culture medium: with embodiment one.

[0155] 2.2. Optimization of VP60 sequence

[0156] According to the insect cell codon preference, the original sequence VP60 of the rabbit hemorrhagic fever virus capsid protein gene measured in Example 1 is optimized without changing the amino acid sequence. The optimized sequence VP60-Y is shown in SEQ ID NO:2, corresponding The amino acid sequence of the amino acid sequence is shown in SEQ ID NO:3. The original sequence contains rare codons in tandem, which reduces the translation sequence and even untranslates the device. The CAI value of the optimized sequence is increased from 0.72 to 0.79, and the GC content and unsuitable peaks are adjusted. In order to extend the half-life of mRNA, the GC content was adjusted from 54.9% to 53....

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Abstract

The invention provides rabbit hemorrhagic disease virus-like particles. The rabbit hemorrhagic disease virus-like particles are prepared by optimizing rabbit hemorrhagic disease virus VP60 gene according to insect cell biased codons, connecting the optimized VP60 gene with an insect cell efficient expression vector, and then producing the rabbit hemorrhagic disease virus-like particles through a rhabdovirus/ insect cell expression system. The rabbit hemorrhagic disease virus-like particles can be mixed with a preservative and adjuvants to prepare a rabbit hemorrhagic disease vaccine. The rabbit hemorrhagic disease virus-like particles can greatly increase the expression quantity of the rabbit hemorrhagic disease virus-like particles, and have the advantages of high safety, high immunogenicity, low cost and the like, and is easy to ferment.

Description

technical field [0001] The invention relates to the field of microorganisms, genetic engineering and veterinary biopharmaceuticals, in particular to a rabbit hemorrhagic disease virus-like particle, its preparation method and application. Background technique [0002] Rabbit hemorrhagic disease, commonly known as rabbit plague, is an acute, virulent, highly contagious and fatal infectious disease caused by rabbit hemorrhagic disease virus (RHDV). Because it was once a devastating infectious disease of rabbits, which brought huge economic losses to the rabbit industry, it has attracted the attention of the rabbit industry. The disease was first reported in China in 1984. In 1989, the World Organization for Animal Health (OIE) officially listed the disease as a Class B infectious disease, and my country listed it as a Class II infectious disease (Wang Yongkun et al., Diagnosis and Prevention of Rabbit Distemper, 1992). [0003] In the seventh report of the International Comm...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/866C12N7/04A61K39/12A61P31/14C12R1/93
Inventor 夏振强郑文文金宏丽张渭蛟
Owner CHANGCHUN SR BIOLOGICAL TECH
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