Gene expression composition, PRRS (Porcine Reproductive and Respiratory Syndrome) oral vaccine and preparation methods thereof

A technology for respiratory syndrome and gene expression, which is applied in the fields of gene expression composition, oral vaccine for porcine reproductive and respiratory syndrome and preparation thereof, can solve the problems of economic loss, increased mortality, serious complications, etc., and avoid safety Effect

Active Publication Date: 2011-03-23
黄鹏林
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

And the younger the pigs, the more serious the complications, le

Method used

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  • Gene expression composition, PRRS (Porcine Reproductive and Respiratory Syndrome) oral vaccine and preparation methods thereof
  • Gene expression composition, PRRS (Porcine Reproductive and Respiratory Syndrome) oral vaccine and preparation methods thereof
  • Gene expression composition, PRRS (Porcine Reproductive and Respiratory Syndrome) oral vaccine and preparation methods thereof

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Experimental program
Comparison scheme
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Embodiment 1

[0064] The construction of embodiment 1 vaccine expression vector

[0065] (丨) Materials and methods

[0066] A. Plastid material

[0067] Plasmid p35PGHT: full-length 8.1kb, containing 0.4kb CaMV 35S promoter, GUS reporter gene, NOS terminator and banana heavy ubiquitin gene 3'MAR.

[0068] Plastid p79PSGHT: 9.1kb in full length, containing the promoter of banana heavy ubiquitin gene (Mh-UBQ1), GUS reporter gene, NOS terminator and banana heavy ubiquitin gene 3'MAR.

[0069] Plastid pGKU: full-length 7.4kb, containing left border and right border, respectively driven by CaMV 35S promoter for selection gene nptII and reporter gene GUS.

[0070] Plastid pBluescript II SK(-): full-length 2.9kb, with multiple cloning site(MCS).

[0071] Among them, plastid pGKU and pBluescript II SK(-) are readily available commercially available plastids; p35PGHT and p79PSGHT were obtained through the following construction strategies:

[0072] Please refer to Taiwan Patent Application (TW p...

Embodiment 2

[0095] The analysis of embodiment 2 expression ORF5 gene tobacco transplantation plant

[0096] Molecular Validation of Tobacco Transplanted Plants

[0097] The result of histochemical staining analysis of GUS activity in the tobacco transgenic plants screened by antibiotics showed a blue positive reaction. Extract genomic DNA from leaves of plants to be transformed, and perform PCR with applicable primers. The results of PCR showed that the pGKU-35PRRSV could be used in the transgenic plants ( Figure 5A ) and pGKU-79PRRSV to be transfected plants ( Figure 5B ) detected the expected synthetic fragment; and the results of Southern hybridization analysis also had the expected signal fragment, in which Figure 5C It is the analysis result of pGKU-35PRRSV to be transformed plants, Figure 5D It is the analysis result of pGKU-79PRRSV to be transformed plants.

[0098] Gene Expression Analysis of Tobacco Transplanted Plants

[0099] Extract the total RNA of tobacco transgeni...

Embodiment 3

[0107] Embodiment 3 expresses the analysis of ORF5 gene banana transplantation plant

[0108] Staining Analysis of Banana Transplanted Plants

[0109] The pGKU-35PRRSV and pGKU-79PRRSV plastids obtained in Example 1 were transformed into bananas, such as: Beijiao (Pei Chiao), and the various parts of the banana transplanted plants were detected with GUS activity histochemical staining to analyze the Whether banana transgenic plants express PRRSV GP5 protein. see Figure 10A to Figure 10D ,The results show, Figure 10A is the root of the control group; Figure 10B Transplant the roots of banana plant seedlings for pGKU-79PRRSV; Figure 10C and Figure 10D Represent the roots and leaves of pGKU-35PRRSV-transformed banana plant seedlings; blue positive reactions can be shown in the roots and leaves, respectively.

[0110] Molecular Verification of Transplanted Banana Plants

[0111] In addition, the genomic DNA of the banana transgenic strain expressing the PRRSV ORF5 gene...

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Abstract

The invention discloses a gene expression composition capable of expressing an allergen protein ORF5 (Open Reading Frame) of a PRRSV (Porcine Reproductive and Respiratory Syndrome Virus), a vaccine expression vector thereof and an oral vaccine containing the same. The gene expression composition comprises a promoter, polynucleotide for coding the ORF5 peptide of the PRRSV, a terminator NOS and polynucleotide of a banana heavy ubiquitin gene 3' MAR. The invention also provides the preparation methods of the gene expression composition, the vaccine expression vector thereof and the oral vaccinecontaining the same. In addition, the invention also provides a method for preventing the PRRS. The vaccine expression vector is transferred into a plant through a transgenic technology, transgenic plants is fed to pigs generate an antibody resisting the PRRSV so as to prevent the PRRS.

Description

technical field [0001] The invention relates to a gene expression composition capable of expressing antigenic protein of porcine reproductive and respiratory syndrome virus, a vaccine expression vector thereof, an oral vaccine containing the gene expression composition and a preparation method thereof. Background technique [0002] Porcine reproductive and respiratory syndrome (PRRS) is a very important porcine viral disease in recent years, causing serious economic losses in Europe, America and all over the world. PRRS is an infectious disease of pigs mainly caused by reproductive disorders of sows and respiratory tract infection of pigs of all ages, and is easily complicated by secondary infection and death. And its complication of younger pig is more serious, causes mortality rate to increase, thereby causes very big loss economically. [0003] The disease first broke out in most pig farms in the United States in 1987. It is also called blue ear disease because the disea...

Claims

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Application Information

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IPC IPC(8): C12N15/11C12N15/63C12N15/29C07K19/00A61K39/12A61P31/14
Inventor 黄鹏林杜宜殷廖育辰林宜佑李盛新黄暐芬
Owner 黄鹏林
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