Extraction method of recombinant human interleukin-2 fermentation inclusion body

An interleukin and extraction method technology, which is applied in the field of extraction of recombinant human interleukin-2 fermented inclusion bodies, can solve the problems of low purity of inclusion bodies, low yield and low expression, and achieves guaranteed culture effect and production saving cost, the effect of improving the recovery rate

Active Publication Date: 2014-08-27
BEIJING FOUR RINGS BIOPHARM
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Problems solved by technology

[0005] However, when the applicant used the above method to replace the engineered bacteria with E.Coli K802 (ply-4) to produce interleukin-2 fermentation inclusion bodies, it was found that the expression level of ril-2 was very low, less than 25%. , resulting in very low purity of the obtained inclusion bodies, which cannot meet the requirements of large-scale production
Then the applicant adjusted the process parameters through the method of

Method used

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Embodiment 1

[0029] The large-scale extraction method for recombinant interleukin 2 of the present invention comprises the steps of: (1) selecting an engineering strain as E.Coli K802 (ply-4) fermentation culture; (2) carrying out high-pressure homogenization and crushing of the bacterial cells obtained from the fermentation culture; (3) Use a high-speed tubular centrifuge to collect crude inclusion bodies, and then wash and refine the collected crude inclusion bodies to obtain high-purity inclusion bodies; (4) Perform denaturation and dissolution on the refined inclusion bodies to obtain inclusion body dissolution liquid.

[0030] The specific technical solutions of the above steps are as follows:

[0031] (1) Select engineering bacteria E.Coli K802 (ply-4) for fermentation culture

[0032] This step further includes seed expansion culture and fermentor culture, and the seed expansion culture includes the following sub-steps:

[0033] a. Strain recovery: Take out a working seed of E.Col...

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Abstract

The invention discloses an extraction method of a recombinant human interleukin-2 fermentation inclusion body. The extraction method comprises the following steps: (1) culturing an engineering strain of an escherichia coli expression recombinant interleukin-2; (2) breaking a thallus obtained by fermentation and culturing to collect an inclusion body; (3) extracting and washing the inclusion body to obtain a refined inclusion body, wherein the engineering strain is E.Coli K802(ply-4), the culturing method comprises multiplication culture and fermentation tank culture, which both adopt a yeast and peptone-containing antibiotic-free culture medium, and the thallus is broken by adopting a high-pressure homogenizer, and is degenerated to obtain a high-purity ril-2 stock solution. The extraction method disclosed by the invention is short in production period, high in production efficiency, large in production scale, and especially suitable for industrialized production; the production cost is lowered.

Description

technical field [0001] The invention relates to the field of biopharmaceuticals, in particular to a method for extracting recombinant human interleukin-2 fermentation inclusion bodies. Background technique [0002] Interleukin-2 (IL-2) is an immunoregulatory factor secreted by T cells, which can not only promote the proliferation of T cells, but also enhance the activity of NK and other immunocompetent cells. , regulation and immune surveillance play an important role. In recent years, it has attracted more and more attention because of its important role in tumor treatment. Since Taniguchi et al. first cloned IL-2 cDNA in 1983 and successfully expressed IL-2 in the monkey cell line COS-7, many scholars have successfully expressed it in E. coli. Since recombinant interleukin-2 (rIL-2) has all the functions of natural IL-2, mass production of IL-2 by genetic engineering will meet the requirements of laboratory and clinical practice. [0003] The biological activity of reco...

Claims

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Application Information

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IPC IPC(8): C07K14/54C07K1/14
CPCC07K14/55
Inventor 程度胜桑建斌吕建龙韩明娣龙应国
Owner BEIJING FOUR RINGS BIOPHARM
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