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Chinese fowlpox virus 282E4 low virulent strain complete genome sequence and application thereof

A technology of genome sequence and fowl pox virus, applied in application, gene therapy, antiviral agent, etc., can solve the problems of limited strain research and application

Inactive Publication Date: 2012-01-18
MILITARY VETERINARY RES INST PLA MILITARY MEDICAL ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] The attenuated strain of FPV282E4 in my country can be used as a safe and effective vaccine to prevent fowlpox. The genetically engineered live carrier vaccines of poultry and mammals prepared with it as a carrier have good application prospects. Up to now, there is no complete genome sequence of the attenuated strain of FPV282E4 in China reports, thus limiting the further research and application of this strain

Method used

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  • Chinese fowlpox virus 282E4 low virulent strain complete genome sequence and application thereof
  • Chinese fowlpox virus 282E4 low virulent strain complete genome sequence and application thereof
  • Chinese fowlpox virus 282E4 low virulent strain complete genome sequence and application thereof

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Embodiment 1

[0022] Example 1 Obtaining the whole genome sequence of Chinese fowlpox virus 282E4 attenuated strain

[0023] 1 Culture of fowl pox virus

[0024] 1.1 Preparation of chicken embryo fibroblasts (CEF)

[0025] 1.1.1 Treatment of chicken embryos

[0026] Take two 9-11-day-old SPF chicken embryos, take them out and put them in a plate under aseptic conditions, remove the head, limbs and viscera, cut them into small pieces and wash them twice with Hank's solution, about 10ml each time. Move the small piece into a 10ml penicillin bottle, cut it into 1-2mm pieces with ophthalmic scissors, add 20ml Hank's solution to wash, let it stand for 2min, make the tissue pieces sink, and the pieces are still in the suspension, then Aspirate the supernatant, and rinse 2-3 times in the same way.

[0027] 1.1.2 Digestion

[0028] Add 4 times the amount of 0.25% trypsin solution (about 2.5ml) to the precipitate, shake and mix well, put it in a water bath at 37°C for 30min, and shake gently onc...

Embodiment 2

[0052] Example 2 Construction and application of novel fowlpox virus vector shuttle plasmid

[0053] Due to the large genome of fowlpox virus (FPV), it is very difficult to directly manipulate genomic DNA, so it is necessary to first construct an FPV expression vector, and then obtain a recombinant virus by homologous recombination. At present, the flanking sequence of TK gene is mostly used as the recombination arm in the construction of recombinant fowlpox virus shuttle expression vector. In addition, the length of the homologous sequence is directly related to the efficiency of recombinant fowlpox virus (rFPV) production. Previous studies have shown that when the length of the homologous sequence is extended from 0.73kb to 4.5kb, the homologous recombination rate can increase linearly from 0.073% to 0.62%. In the case of unknown genome background of FPV, it is difficult to obtain the sequences on both sides of the TK gene, which has caused certain difficulties for the cons...

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Abstract

The invention for the first time provides a Chinese fowlpox virus 282E4 low virulent strain complete genome sequence shown as a SEQ ID No.1. The sequence can be applied to researches on fowlpox virus functional genomics, novel viral vector construction and fowlpox virus prophylactic vaccine. The sequence provides objective basis for further clarifying genetic derivation characteristics and genome background of Chinese fowlpox virus 282E4 low virulent strain, provides reliable data for further developing novel fowlpox virus prophylactic vaccine and rapid diagnosis and detection reagent, and lays foundations for further studying hereditary variation rules, pathogenic mechanism, infection mechanism and immunization mechanism of fowlpox virus.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to the whole genome sequence of the Chinese chickenpox virus 282E4 attenuated strain and its application. Background technique [0002] Fowlpox virus (FPV) belongs to the genus Fowlpoxvirus of the family Poxviridae. The mature virus particles are brick-shaped or ellipsoidal in size, with a size of 280nm×330nm and a mulberry-like surface. It is the largest animal virus found so far. One of the largest and most complex viruses. [0003] FPV is a representative of fowlpox virus among poxviruses. Compared with other poxviruses, FPV has a larger genome and can accommodate more foreign genes without losing its infectivity; foreign proteins can be faithfully expressed in host cells and Carry out necessary glycosylation, carboxylation and other modifications; the expression product has good immunogenicity and can induce long-term cellular immunity and humoral immunity in the body; ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/39C12N15/863C12N15/63C12N1/15C12N1/19C12N1/21C12N5/10A61K48/00A61P31/20G01N33/569
Inventor 金宁一李昌刘存霞李霄杜寿文王茂鹏朱娜孙丹丹秦艳青李沂鲁会军田明尧金扩世
Owner MILITARY VETERINARY RES INST PLA MILITARY MEDICAL ACAD OF SCI
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