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Recombinant Newcastle disease heat-resistant vaccine strain for expression of signal peptide-replaced H5 subtype avian influenza virus HA protein and preparation method thereof

A signal peptide and avian influenza technology, applied in the field of molecular biology, can solve the problems of inability to prepare live vaccines, the risk of dispersing the virus, and the high cost of immunization, so as to improve the thermal stability of the vaccine, enhance the secretion expression and immunogenicity, The effect of remarkable heat resistance characteristics

Inactive Publication Date: 2016-11-09
INST OF ANIMAL SCI & VETERINARY HUBEI ACADEMY OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The existing H5 avian influenza and Newcastle disease dual vaccines that have been on the market are all prepared after inactivating the H5 subtype avian influenza virus strain and the Newcastle disease virus strain. The main reason is that the H5 subtype avian influenza virus has a certain pathogenicity and cannot be used in the preparation of live vaccines
The disadvantage of this type of vaccine is that it requires intramuscular injection, which is labor-intensive and time-consuming, and the cost of immunization is high
The use of inactivated vaccines has increased the difficulty of clinically distinguishing wild virus infection from vaccine immunity to a certain extent, and there is a risk of spreading the virus

Method used

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  • Recombinant Newcastle disease heat-resistant vaccine strain for expression of signal peptide-replaced H5 subtype avian influenza virus HA protein and preparation method thereof
  • Recombinant Newcastle disease heat-resistant vaccine strain for expression of signal peptide-replaced H5 subtype avian influenza virus HA protein and preparation method thereof

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0025] The first step, the construction and identification of the recombinant full-length plasmid pTS-tHA / H5

[0026] 1.1 PCR amplification of HA gene of H5 subtype avian influenza virus

[0027] The H5N1 avian influenza virus A / chicken / Hubei / 489 / 2004 strain was propagated on 9-11 days old chicken embryos, and the allantoic fluid of the chicken embryos was harvested 5 days after inoculation. The harvested allantoic fluid was subjected to RT-PCR amplification of the HA gene. The amplification primers were: upstream primer 5'-AGAGAGGGCTCTGCTGTGTGCTGCTGCTGTGTGGAGCAGTCTTCGTTTCGCCCAGCGAAATCTCTTCAATTTGCATTGGTTACCATGC-3', downstream primer 5'-ATCGGGGCACTCCGATTCTACCCGTATTTTTTCTUS-ATTATTATTATCCTInCTTTTTTC-3' for cloning (underlined linked complementary sequences). The target band detected by agarose gel electrophoresis is about 1.1kb, which is consistent with the expected 1101bp (the length of the HA1 gene that deletes its own signal peptide is 987bp, and the gene start sequence and g...

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Abstract

The invention discloses a recombinant Newcastle disease heat-resistant vaccine strain for expression of a signal peptide-replaced H5 subtype avian influenza virus HA protein and a preparation method thereof. The heat-resistant vaccine strain is named as rTS-tHA / H5, is preserved in the Wuhan university China center for type culture collection on April 19, 2016 and has an accession number of CCTCC NO: V201628. After infection of animals or cells with the vaccine strain, the vaccine strain can highly express a H5 subtype avian influenza virus truncated HA protein and the tPAs of the vaccine strain can improve secretory expression and immunogenicity of the truncated HA protein. The vaccine strain can be used for preparation of a chicken Newcastle disease and H5 subtype avian influenza bivalent heat-resistant living vaccine.

Description

technical field [0001] The invention belongs to the field of molecular biology, and specifically relates to a recombinant Newcastle disease heat-resistant vaccine strain expressing signal peptide-substituted H5 subtype avian influenza HA protein and a preparation method thereof. Background technique [0002] H5 subtype avian influenza virus can cause an acute fatal infectious disease of poultry - highly pathogenic avian influenza, its clinical manifestations are high mortality, decreased egg production, and severe respiratory symptoms. H5 subtype avian influenza can also infect humans and cause disease. Inoculation of inactivated vaccines is the main means of prevention and control of H5 subtype avian influenza in my country. Traditional inactivated vaccines are relatively simple to prepare, convenient to store and transport, and have long-lasting immune effects. They have played an important role in the prevention and control of avian influenza. However, inactivated vacci...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N7/01C12N15/85A61K39/17A61K39/145A61P31/14A61P31/16C12R1/93
CPCA61K39/12A61K2039/70C12N7/00C12N15/85C12N2760/16134C12N2760/18121C12N2760/18134C12N2760/18151C12N2800/107A61K2300/00
Inventor 邵华斌温国元罗青平潘兹书乔磊罗玲王红琳张腾飞汪宏才张蓉蓉卢琴艾地云
Owner INST OF ANIMAL SCI & VETERINARY HUBEI ACADEMY OF AGRI SCI
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