Preparation method and application of carp spermary bioreactor

A bioreactor and carp technology, applied in the biological field, can solve problems such as complex biochemical preparation process, virulence recovery, environment and production personnel health threats, and achieve the effect of fast growth and large egg production

Inactive Publication Date: 2013-09-04
WUHAN DABANG BIOLOGICAL TECH
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  • Claims
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AI Technical Summary

Problems solved by technology

The production of whole-virus inactivated vaccines has potential safety hazards, which may pose a threat to the environment and the health of production personnel; the development of live attenuated vaccines is difficult, and there is a potential risk of virulence recovery and the formation of new influenza strains
Traditional subunit vaccines are obtained from virus particles. Although the safety is guaranteed, the biochemical preparation process is complex and costly, making it difficult to popularize and apply

Method used

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  • Preparation method and application of carp spermary bioreactor
  • Preparation method and application of carp spermary bioreactor
  • Preparation method and application of carp spermary bioreactor

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0039] A preparation method of a carp testis bioreactor is prepared through the following steps:

[0040] 1) Screening the promoters of highly expressed genes in semen: SDS-PAGE electrophoresis separation of zebrafish semen proteins, cutting the electrophoresis bands for mass spectrometry detection, and obtaining six genes specifically expressed in zebrafish semen; extracting zebrafish semen Testis total RNA, RT-PCR screened out the two genes with the highest expression levels among the six genes, the sequence numbers of the gene bank are BC122153 (SEQ ID NO.1) and BC076027 (SEQ ID NO.3); and the Zebrafish Genome Database Compare and obtain the promoter sequences of these two genes, named as Q2, whose nucleotide sequence is shown in SEQ ID NO.2; Q3, whose nucleotide sequence is shown in SEQ ID NO.4;

[0041] 2) Construction of recombinant vectors: construct gene expression vectors for target protein genes driven by Q2 and Q3 respectively by overlapping PCR method;

[0042] 3)...

Embodiment 2

[0046] A kind of application of fish testis bioreactor in preparing H5N1-HA, its steps are:

[0047] A. the screening of testis specific highly expressed gene promoter, its steps are:

[0048] Firstly, the semen protein of zebrafish was separated by SDS-PAGE electrophoresis, the electrophoresis band was cut for mass spectrometry detection, and the mass spectrometry detection results were analyzed by bioinformatics to obtain six genes specifically expressed in zebrafish semen (Table 1);

[0049] Then total RNA was extracted from zebrafish testis, and cDNA was obtained by reverse transcription. RT-PCR verification (see Table 2 for RT-PCR primers) the expression of the above six genes in the testis ( figure 1 );

[0050] Finally, the two genes with the highest expression levels were selected, and the sequence numbers were BC122153, whose nucleotide sequence was shown in SEQ ID NO.1, and BC076027, whose nucleotide sequence was shown in SEQ ID NO.3, and zebrafish Comparing the ...

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Abstract

The invention discloses a preparation method and application of a carp spermary bioreactor. The preparation method comprises the following steps: A, screening a promoter overexpressing gene in a seminal fluid; B, constructing a target gene expression vector specifically expressed in fish spermary; C, introducing a recombinant gene segment into a red carp oosperm to stably ingrate the recombinant gene segment in genome; and D, letting the target gene efficiency expresse in the transgenic red carp spermary to obtain a bioreactor highly expressing the target gent. According to the invention, by utilizing the property of fish spermary gene specific expression, the bioreactor with safety, high efficiency, and sustainable utilization can be constructed; simultaneously fish have the characteristics of fast speed of enlarging quantity and low breeding cost, thus large-scale low-cost operation of the bioreactor can be realized.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a preparation method of a fish testis bioreactor, and also relates to an application of a carp testis bioreactor in preparing medicinal proteins, industrial enzymes, vaccines and antibodies. Background technique [0002] Transgenic bioreactor refers to the transfer of exogenous genes into cells, animals, plants, and microorganisms, and the use of cell proliferation or organism metabolism to prepare expression products of exogenous genes, mainly including transgenic microbial bioreactors, transgenic animal bioreactors and transgenic plants Three types of bioreactors. [0003] The products produced by transgenic microbial bioreactors generally do not have biological activity, and must undergo a series of modification processes such as glycosylation and hydroxylation, which are cumbersome and time-consuming. The products of transgenic animal and plant bioreactors have natur...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12N15/85A01K67/027C07K14/11C12N9/00A61K39/00C07K16/00
Inventor 阮慧云李良明
Owner WUHAN DABANG BIOLOGICAL TECH
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