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A kind of fermentation method for large-scale production of recombinant exendin-4 polypeptide

A fermentation method and large-scale technology, which is applied in the field of fermentation for large-scale production of recombinant Exendin-4 polypeptides, and can solve problems such as toxic substance residues

Active Publication Date: 2022-07-29
BEIJING GENETECH PHARML
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Problems solved by technology

[0003] Therefore, the technical problem to be solved in the present invention is to overcome the defect of toxic substances remaining in the fermentation production of recombinant Exendin-4 polypeptide in the prior art, thereby providing a fermentation method for producing recombinant Exendin-4 polypeptide, which will not produce toxicity residual substance

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  • A kind of fermentation method for large-scale production of recombinant exendin-4 polypeptide
  • A kind of fermentation method for large-scale production of recombinant exendin-4 polypeptide
  • A kind of fermentation method for large-scale production of recombinant exendin-4 polypeptide

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Embodiment 1

[0035] Example 1 Preparation of recombinant Exendin-4 polypeptide

[0036] See the overall process flow Figure 7 , In this example, the induction concentration, induction temperature and induction time of lactose were determined through experimental exploration. Using lactose as an inducer, the expression levels of TB, LB and LM9 in three media were compared. After determining the choice of TB medium, the effects of lactose addition on fermentation and expression were compared. Scale-up and research are carried out in fermenters based on groping experimental data.

[0037] 1. Determination of lactose-inducing concentration

[0038] The bacteria were inoculated into 250mL shake flasks (containing 50mL LB medium), and cultivated to OD at 37°C 600 Value 2, add lactose according to 5, 10, 20, 30, 40g / L (final concentration), inductive culture for 4 hours, take samples for testing, compare the effects of different lactose concentrations on expression, the results show that whe...

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Abstract

The present invention provides a fermentation method for large-scale production of recombinant Exendin-4 polypeptide, which adopts pET32a(+)-Exendin-4 / BL21(DE3) fermentation culture, and uses lactose as an inducer for fermentation culture during fermentation. Lactose replaces IPTG as the inducer for the fermentation and culture of engineering bacteria. During the fermentation process, lactose will be catabolized by the bacteria as a carbon source, and no toxic residues will be produced. In addition, in order to make the process of recombinant Exendin‑4 polypeptide more suitable for industrial The fermentation process parameters were optimized, the fermentation scale was expanded from 30L to 500L, and the optimized bacterial body wet weight and protein expression (up to 14.1%-14.6%) were better than the previous process.

Description

technical field [0001] The invention relates to the field of product fermentation, in particular to a fermentation method for large-scale production of recombinant Exendin-4 polypeptide. Background technique [0002] As an inducer for genetically engineered bacteria to regulate the lactose operon, IPTG can activate the lactose operon and express the gene encoding the rExendin-4 fusion protein downstream of the operon, thereby obtaining a fusion protein containing rExendin-4. IPTG will not be degraded during the culture of engineered bacteria, and IPTG has potential toxicity, which affects the safety of the final stock solution. SUMMARY OF THE INVENTION [0003] Therefore, the technical problem to be solved by the present invention is to overcome the defect of the residual toxic substances in the fermentation production of recombinant Exendin-4 polypeptide in the prior art, thereby providing a fermentation method for producing recombinant Exendin-4 polypeptide, which does n...

Claims

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Application Information

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IPC IPC(8): C12P21/02C07K14/575C12R1/19
CPCC12P21/02C07K14/57563
Inventor 樊欣迎李静郭静雅刘月峰梁国龙闻亚磊
Owner BEIJING GENETECH PHARML
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