Gene and method for preparing recombination fugu rubripe interleukin-2 protein

A technology of puffer and interleukin, applied in botany equipment and methods, biochemical equipment and methods, genetic engineering, etc., to achieve high repeatability, simple and convenient operation, and high-efficiency expression

Inactive Publication Date: 2017-02-22
DALIAN OCEAN UNIV
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  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, so far, there is no relevant report on prokaryotic expression an

Method used

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  • Gene and method for preparing recombination fugu rubripe interleukin-2 protein
  • Gene and method for preparing recombination fugu rubripe interleukin-2 protein
  • Gene and method for preparing recombination fugu rubripe interleukin-2 protein

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Embodiment Construction

[0019] The present invention optimizes codons based on the natural redfin puffer interleukin 2 gene for the codon preference of Escherichia coli, and adds enzyme cutting sites and tag sequences at both ends of the sequence. The optimized gene DNA sequence is as SEQ IDNO: 1, used to prepare recombinant red fin puffer interleukin 2 protein.

[0020] The preparation method of recombinant red fin puffer interleukin 2 protein of the present invention, carries out according to the following steps successively:

[0021] a. Synthesize the gene shown in SEQ ID NO:1 and clone the gene shown in SEQ ID NO:1 into the prokaryotic expression vector pET-32a, activate TOP10 / pET-32a / IL-2 cells, and extract the prokaryotic expression plasmid pET-32a / IL-2;

[0022] b. Transform the prokaryotic expression plasmid pET-32a / IL-2 into Escherichia coli BL21(DE3), screen positive clones, perform PCR identification, and then digest with XhoI and NcoI ( figure 1 ) for further identification and construc...

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Abstract

The invention discloses a gene capable of being efficiently expressed for preparing recombination fugu rubripe interleukin-2 and a preparation method. The expression product of the gene has the obvious biological activity. The method comprises the following steps that the gene shown in SEQ ID NO:1 is cloned to a prokaryotic expression vector pET-32a, and prokaryotic expression plasmid is constructed; the prokaryotic expression plasmid is transformed into escherichia coli BL21(DE3), positive clones are screened out, and recombinant escherichia coli BL21(DE3) is constructed; LB is used as a basic culture medium, and amplification culture is performed on the recombinant escherichia coli BL21(DE3); an IPTG inductive agent is added into an amplification culture for induction culture, and thalli is collected through centrifuging; ultrasonication is performed on the collected thalli, a supernate is taken after centrifuging is performed, and a crude recombination fugu rubripe interleukin-2 protein product is obtained.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a gene and method for preparing recombinant redfin puffer interleukin-2 protein which can be expressed efficiently and the expression product has obvious biological activity. Background technique [0002] Redfin puffer is the main species of fish cultured in China, but there are often some viral and bacterial diseases in the culture, such as: white mouth disease, red sea bream iridescent virus disease, vibriosis, streptococcal disease and so on. For these diseases, the solution that often comes to mind is to use chemical drugs or antibiotics, but this method has many problems. In recent years, natural immune products have played an important role in this regard, which can replace antibiotics as a safe drugs are used in practice. [0003] Interleukins (IL) for short, are a family of cytokines that can interact between immune cells and leukocytes, and interleukin-2 (IL-2) ...

Claims

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Application Information

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IPC IPC(8): C12N15/26C12N15/70C07K14/55C07K1/22
CPCC07K14/55C12N15/70C12N2800/101C12N2800/22
Inventor 王秀利孙鹤李慧孔德荣刘海映仇雪梅
Owner DALIAN OCEAN UNIV
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