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Porcine rotavirus recombinant protein, recombinant adenovirus expressing same protein and application

A porcine rotavirus, recombinant adenovirus technology, applied in applications, viruses, viral peptides, etc., can solve the problems of no human adenovirus vector, low probability of repeated use of vaccines, and achieve low dosage and high yield. Effect

Active Publication Date: 2021-04-09
WUHAN KEQIAN BIOLOGY CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Although the application of human adenovirus vector vaccines in humans still needs more in-depth research, it is very promising as an animal vaccine because there is no neutralizing antibody to human adenovirus vectors in animals, and the probability of repeated use of the same vector vaccine is also low. very low

Method used

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  • Porcine rotavirus recombinant protein, recombinant adenovirus expressing same protein and application
  • Porcine rotavirus recombinant protein, recombinant adenovirus expressing same protein and application
  • Porcine rotavirus recombinant protein, recombinant adenovirus expressing same protein and application

Examples

Experimental program
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Effect test

Embodiment 1

[0035] This embodiment provides a method for constructing a recombinant adenovirus, the specific process is as follows:

[0036] 1. Construction of transfer vector pShuttle-CMV-G5VP7-G9VP8

[0037] 1.1 Obtaining the target genes PoRV-VP7 and VP8

[0038] According to the codon preference of adenovirus expression system, the nucleotide sequences of porcine rotavirus type 5 VP7 protein and type 9 VP8 protein were codon optimized. Gene synthesis was performed by Sangon Bioengineering Co., Ltd. The synthetic gene sequence is shown in SEQ ID NO: 1 (also contains an IL-2 signal peptide at the beginning). The synthetic target gene is placed on the plasmid vector pUC-SP, which is obtained in the form of pUC-SP-G5VP7-G9VP8.

[0039] 1.2 Construction of recombinant transfer vector pShuttle-CMV-PoRV-G5VP7-G9VP8

[0040] Insert the G5-VP7 and G9-VP8 genes of PoRV into the pShuttle-CMV vector by recombination, see the attached map figure 1 :

[0041] (1) PCR amplification: through PC...

Embodiment 2

[0070] In this example, the recombinant adenovirus prepared in Example 1 was used to prepare a vaccine, and its performance was tested. The process is as follows:

[0071] 1. Evaluation of adenovirus Ad-PoRV-G5VP7-G9VP8 vector vaccine in pigs

[0072] 1.1 Safety evaluation of adenovirus Ad-PoRV-G5VP7-G9VP8 vector vaccine in pigs: 8 experimental pigs (PoRV-negative pigs aged 14-21 days) were purchased and divided into two groups, A and B, with 4 pigs in each group . Group A intramuscular injection of 10 per head and neck 10 TCID 50 Recombinant adenovirus (1mL); group B was intramuscularly injected with 1mL sterilized PBS on the head and neck, and observed continuously for 14 days. The pigs in the two groups were in the same state and had no abnormal reaction, which indicated that the vaccine was safe for pigs.

[0073] 1.2 Immunogenicity evaluation of adenovirus Ad-PoRV-G5VP7-G9VP8 vector vaccine in pigs: 12 experimental pigs were purchased and randomly divided into three gr...

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Abstract

The invention relates to the technical field of veterinary biology, and particularly relates to a porcine rotavirus recombinant protein, a recombinant adenovirus expressing same protein and application. The recombinant protein is formed by connecting a porcine rotavirus type 5 VP7 protein and a porcine rotavirus type 9 VP8 protein, and specifically comprises an amino acid sequence shown as SEQ ID NO: 1. Gene sequences of coding genes of the porcine rotavirus type 5 VP7 protein and the porcine rotavirus type 9 VP8 protein are constructed into an adenovirus expression vector, codon optimization is carried out, and the recombinant adenovirus for the porcine rotavirus is prepared. The structure and function of the target protein expressed by the recombinant adenovirus are completely consistent with those of the natural state, and the recombinant adenovirus has high safety and immunogenicity.

Description

technical field [0001] The invention relates to the technical field of veterinary medicine, in particular to a porcine rotavirus recombinant protein, a recombinant adenovirus expressing the protein and application thereof. Background technique [0002] Porcine Rotavirus (RoRV) is a double-stranded RNA virus without an envelope, belonging to the genus Rotavirus of the Reoviridae family. As one of the important pathogens, the harm of porcine rotavirus to the pig industry mainly lies in causing viral diarrhea in piglets. Piglet viral diarrhea caused by porcine rotavirus infection refers to a disease of piglets caused by porcine rotavirus with vomiting, anorexia, dehydration, diarrhea and acid-base balance disorder as the main symptoms. The viral infection caused by porcine rotavirus is common in pig herds no matter it is single infection or cross-infection, resulting in the sickness or even death of piglets, which has caused huge economic losses to the pig breeding industry. ...

Claims

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Application Information

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IPC IPC(8): C07K14/14C12N15/46C12N15/861A61K39/15A61K39/295A61P31/14
CPCC07K14/005C12N15/86A61K39/12A61P31/14C12N2720/12322C12N2710/10043C12N2720/12334A61K2039/552A61K2039/70
Inventor 张华伟徐高原陈映瑾周明光罗修鑫周飞燕郝根喜汤细彪
Owner WUHAN KEQIAN BIOLOGY CO LTD
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