Newcastle disease virus-like particles, preparation method and application thereof

A technology of Newcastle disease virus and particles, which is applied in the field of Newcastle disease vaccine, can solve the problems of high cost, environmental pollution, and small immune adverse reactions, and achieve the effects of increased yield and purity, good immune effect, and good industrial application prospects

Active Publication Date: 2017-05-31
JILIN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Benefits of technology

This new method described for producing an infectious agent that contains viruses with similar properties but different structures allows them to be more effective at stimulating antibody production against other pathogens or tumors than existing methods like live bacteria.

Problems solved by technology

The technical problem addressed in this patents relates to developing novel methods for generating effective anti-influenza agents against Newcastle Diseas through molecules derived from their own components called ribon shell/nucleate polymer(RNA)/lipid complexes.

Method used

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  • Newcastle disease virus-like particles, preparation method and application thereof
  • Newcastle disease virus-like particles, preparation method and application thereof
  • Newcastle disease virus-like particles, preparation method and application thereof

Examples

Experimental program
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Embodiment 1

[0053] The Newcastle disease virus-like particle provided by the present invention is a protein particle with a hollow structure formed by self-assembly of M protein, F protein, NP protein and HN protein; wherein the base sequence of the M1 gene corresponding to the M protein is as shown in SEQ ID NO .1, the base sequence of the F1 gene corresponding to the F protein is shown in SEQ ID NO.2, the base sequence of the NP1 gene corresponding to the NP protein is shown in SEQ ID NO.3, and the base sequence of the HN1 gene corresponding to the HN protein The base sequence is shown in SEQ ID NO.4.

[0054] What needs to be explained is that when preparing NDV-like particles, the method of genetic engineering is generally adopted, that is, the specific gene sequence is integrated into the genome of the host cell through genetic engineering technology, and then expressed in the host cell, and finally Purified protein particle samples are obtained through specific separation and purifi...

Embodiment 2

[0056] This example briefly introduces the construction process of the recombinant baculovirus expressing the structural protein of Newcastle disease virus in the preparation process of Newcastle disease virus-like particles as follows.

[0057] (1) Amplify the sequence of each structural protein gene

[0058] According to the target base sequence shown in SEQ ID NO.1~4 in Example 1, referring to the prior art, M1 gene, F1 gene, NP1 gene, HN1 gene were obtained by artificial amplification by polymerase chain reaction (PCR method) base sequence.

[0059] The primer sequences for PCR amplification are specifically:

[0060] M1 upstream primer: 5'-ATGGACTCATCTAGGACTATTGGACT-3',

[0061] M1 downstream primer: 5'-TTATTTACGGAAGGGGTTGTATTTAGC-3';

[0062] F1 upstream primer: 5'-ATGGGTTCGAAGCCATCCACCCG-3',

[0063] F1 downstream primer: 5'-TTAGGCTCTCGTTGTAGCCCT-3';

[0064] NP1 upstream primer: 5'-ATGTCCAGCGTCTTCGACG-3',

[0065] NP1 downstream primer: 5'-TTAGTAACCCCAGTCAGTATCGT...

Embodiment 3

[0108] This example briefly introduces the expression, assembly and purification process of structural proteins in NDV-like particles as follows.

[0109] (1) Expression of structural proteins in Newcastle disease virus-like particles

[0110] The supernatant containing the fourth-generation recombinant baculovirus rBV-M1, rBV-F1, rBV-NP1, rBV-HN1 collected in Example 2 was added to the suspension culture at a volume ratio of 3:2:1:2 Insect Sf9 cells (3mL of rBV-M1, 2mL of rBV-F1, 1mL of rBV-NP1, 2mL of rBV-HN1 were added to the total volume of 300mL cell culture medium), and the infection time was 96 hours;

[0111] After infection, the infected cells were cultured on a shaker at 28°C and 120r / min;

[0112] During this culture process, the structural proteins self-assemble after being expressed separately in host cells, and the assembled virus-like particles are secreted into the cell culture supernatant.

[0113] (2) Purification of Newcastle disease virus-like particles ...

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Abstract

The invention relates to the technical field of Newcastle disease vaccine in particular to Newcastle disease virus-like particles, a preparation method and a patent application of an application thereof. The virus-like particles are protein particles with a hollow structure formed by the self-assembly of Newcastle disease virus M protein, F protein, NP protein and HN protein. The base sequence of a M1 gene corresponding to the M protein is shown in SEQ ID NO.1, the base sequence of the F1 gene corresponding to the F protein is shown in SEQ ID NO.2, the base sequence of the NP1 gene corresponding to the NP protein is shown in SEQ ID NO.3, the base sequence of the HN1 gene corresponding to the HN protein is shown in SEQ ID NO. 4. The Newcastle disease virus-like particles, preparation method and application have the technical advantages of high expression level of structural protein, self-assembly of the virus-like particles, immunogenicity closer to real virus particles, high yield and purity of the virus-like particles, and good immunization effect.

Description

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Claims

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Application Information

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Owner JILIN UNIV
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