Fusion gene containing transmissible gastroenteritis virus (TGEV) and porcine epidemic diarrhea virus (PEDV) protective antigens as well as encoding protein and application thereof

A technology of fusion gene and protein, applied in the fields of fusion gene, encoded protein and application

Inactive Publication Date: 2015-07-29
BEIJING ACADEMY OF AGRICULTURE & FORESTRY SCIENCES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the development of transgenic plant vaccines by combining TGEV and PEDV protective antigen fragments in tandem has not

Method used

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  • Fusion gene containing transmissible gastroenteritis virus (TGEV) and porcine epidemic diarrhea virus (PEDV) protective antigens as well as encoding protein and application thereof
  • Fusion gene containing transmissible gastroenteritis virus (TGEV) and porcine epidemic diarrhea virus (PEDV) protective antigens as well as encoding protein and application thereof
  • Fusion gene containing transmissible gastroenteritis virus (TGEV) and porcine epidemic diarrhea virus (PEDV) protective antigens as well as encoding protein and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0033] The preparation method of the TGEV CB antigenic site and PEDV COE antigenic site tag protein expressed and purified by the Escherichia coli used in Example 1

[0034] (1) First obtain the cDNA sequences of TGEV and PEDV in the GenBank of the NCBI website, the codes are GenBank accession No.AJ271965.2 and No.JN825712.1, and the base sequences of the S-encoded genes are located at 20365-24708 (TGEV , a total of 4344bp) and 20634-24794 (PEDV, a total of 4161bp); select the 118-510 (including the CB antigenic site) in the S coding gene in TGEV and the 1363-1968 (including the COE antigenic site) fragment in the S coding gene in PEDV Cloned into the prokaryotic expression vector pET-32a (purchased from Novagen), identified by enzyme digestion and sequencing, and transformed the positive plasmids (pET-32a-CB and pET-32a-COE) into competent cells of the expression strain Rosetta (DE3) (purchased from Kangwei Century Biotechnology Co., Ltd.) for expression, construction referen...

Embodiment 2

[0038] Embodiment 2 Containing the acquisition of the fusion gene sTP of TGEV and PEDV protective antigen

[0039] Firstly, obtain the cDNA sequence of TGEV in GenBank of NCBI website, coded as GenBank accession No.AJ271965.2, in which the base sequence of the S coding gene is located at 20365-24708, a total of 4344bp, according to the existing relevant TGEV protective antigenic sites In the study, the base sequences of 118-510 (CB antigenic site), 805-1239 (D antigenic site) and 1537-1986 (A antigenic site) in the S coding gene of TGEV were selected as the basic sequence. Then obtain the cDNA sequence of PEDV in the GenBank of the NCBI website, coded as GenBank accession No. JN825712.1, wherein the base sequence of the S coding gene is located at 20634-24794, a total of 4161bp; according to the existing relevant PEDV protective antigenic sites In the study, the base sequences of 1363-1968 (COE antigenic site), 2221-2340 (Be epitope antigenic site) and 4084-4158 (P1 antigenic ...

Embodiment 3

[0040] Example 3 Construction of recombinant expression vector pBAC9090

[0041] (1) Construction of plant expression vector pBAC198, the construction method is as follows:

[0042] The plasmid vector pBI221 (purchased from Clontech, USA) was double-digested with SacI and EcoRI to obtain the 0.26kb terminator sequence of the NOS3' terminal of the Agrobacterium tumefaciens nopaline synthase gene, and then the NOS3' of the Agrobacterium tumefaciens nopaline synthase gene The terminator sequence was inserted into the SacI and EcoRI sites of the basic plasmid pSP72 (purchased from Promega), and the correctly connected plasmid was obtained by blue-white screening method, which was named pBPC18 (2.7kb).

[0043] The Zein promoter (1435bp, base sequence as shown in SEQ ID NO.3) is an artificially synthesized sequence, synthesized by Shanghai Sangon Company. The Zein promoter has a HindIII restriction site at the 5' end and a BamHI restriction site at the 3' end. The Zein promoter is...

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Abstract

The invention discloses a fusion gene containing transmissible gastroenteritis virus (TGEV) and porcine epidemic diarrhea virus (PEDV) protective antigens as well as encoding protein and application of the fusion gene. The base sequence of the fusion gene containing the TGEV and the PEDV protective antigens is shown by SEQ ID No. 1. The protein for encoding the fusion gene has the amino acid sequence shown by SEQ ID No. 2. The fusion gene is introduced into corns, and transgenic corn plants containing the TGEV and the PEDV protective antigens are obtained by screening and indentifying. The transgenic corn vaccine is capable of efficiently expressing the fusion protein containing the TGEV and the PEDV protective antigens, and the expression product has remarkable immunogenicity.

Description

technical field [0001] The invention relates to a fusion gene, encoded protein and application, in particular to a fusion gene containing TGEV and PEDV protective antigen, encoded protein and application. Background technique [0002] Porcine transmissible gastroenteritis is characterized by vomiting, severe diarrhea, dehydration and high lethality to piglets under 2 weeks of age. Pigs of different breeds and ages are susceptible. The disease is caused by porcine transmissible gastroenteritis virus (Transmissible Gastroenteritis Virus (TGEV), which mainly harms newborn piglets. As the age increases, the mortality rate of the disease gradually decreases, but it often causes production performance decline, feed return rate reduction, and greater economic losses. Porcine epidemic diarrhea disease is characterized by diarrhea, vomiting and dehydration. Pigs of all ages and breeds are susceptible, especially suckling piglets. The disease is caused by porcine epidemic diarrhea vir...

Claims

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Application Information

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IPC IPC(8): C12N15/62C12N15/82C07K19/00A01H5/00
Inventor 王金洛张晓东周宏专杨兵薛静徐福洲张立全
Owner BEIJING ACADEMY OF AGRICULTURE & FORESTRY SCIENCES
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