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Preparation method of human epidermal growth factor

An epidermal growth factor and human production technology, applied in the field of bioengineering, can solve the problems of complex extraction process, high operating cost, and no fermentation production process yet.

Inactive Publication Date: 2011-10-12
孙民富
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] Although there are some genetic engineering production methods of EGF at present, these methods all have the disadvantages of low product concentration in the fermentation broth, complex extraction process, high operating cost and low yield. Therefore, so far, there is no mature industrialization Fermentation Production Process

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0017] (1) Engineering bacteria fermentation:

[0018] Pick a single colony from the strain tube stored at -70°C and insert it into 5-15ml seed medium, add ampicillin at the same time to make the concentration in the medium reach 50-200mg / ml, 28-32°C, 150 -250rpm shaker culture for 8-12 hours, according to 1-8% inoculum size, the previous culture solution was reintroduced into the seed medium, and ampicillin was added at the same time to make it at 50-200mg / ml, 28-32 ° C, 150 -250rpm shaker culture for 8-12 hours, then add the activated seed liquid to the fermentation medium according to the inoculation amount of 1-8%, 29-31 ° C, 160-240rpm, maintain the pH value at 7.0, and cultivate for 9-11 hours, Then the temperature was induced and the cultivation was continued for 2-8 hours. Human epidermal growth factor (EGF) is expressed in the strain.

[0019] (2) Expanded bed adsorption:

[0020] Use a refrigerated centrifuge to centrifuge the fermentation broth at a speed of 6000...

Embodiment 2

[0026] (1) Engineering bacteria fermentation:

[0027] Pick a single colony from the strain tube stored at -70°C and insert it into 10ml of seed medium, add ampicillin at the same time to make the concentration in the medium reach 100mg / ml, culture at 30°C, 200rpm shaker for 10 hours, Insert the previous culture solution into the seed medium again by 5% inoculum amount, add ampicillin simultaneously, make it at 100mg / ml, 30 ℃, 200rpm shaker culture 10 hours, press the activated seed again by 5% inoculum amount The solution was added to the fermentation medium, maintained at 30° C., 200 rpm, and the pH value was 7.0, and cultivated for 10 hours, then induced by temperature, and continued to cultivate for 4 hours. Human epidermal growth factor (EGF) is expressed in the strain.

[0028] (2) Expanded bed adsorption:

[0029] Use a refrigerated centrifuge to centrifuge the fermentation broth at a speed of 10,000 rpm for 30 min at 4°C, discard the supernatant and save the precipit...

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Abstract

The invention discloses a preparation method of human epidermal growth factor (EGF). The method comprises the following steps: fermenting with engineering bacteria, picking up a single colony from a preserved bacteria tube to inoculate in seed culture medium, adding Ampicillin, culturing with a shaker; performing expanded bed adsorption, using a refrigeration centrifuge to centrifuge the fermentation solution, discarding fermentation liquid to reserve precipitate, performing ultrasonic processing, centrifuging, adding the product in an expanded bed, eluting, collecting the elution peak with EGF, reserving the elution solution to store for standby; performing gel separation, using a chromatography system to add the elution solution in a gel chromatographic column, buffering the elution solution, reserving the absorption peak; and adopting the sublimation freezing-drying technology to obtain the finished product. The invention has the advantages of simple method, high product concentration, low operating cost, high yield and simple extraction technology.

Description

field of invention [0001] The invention relates to a preparation method for producing human epidermal growth factor (EGF), which belongs to the field of bioengineering. Background of the invention [0002] In 1962, Dr. Stanley Cohen of the United States and Professor Rita Levi-Montalcini discovered the "epidermal growth factor" EGF (Epidermal Growth Factor) that determines the age of the skin, and they won one of the highest scientific awards in the world—— Nobel Prize in Physiology. [0003] Epidermal growth factor is an active substance in the human body. It is a living cell composed of 53 amino groups. By stimulating the tyrosine phosphorylation of the epidermal growth factor receptor, it can repair and proliferate the surface cells of the skin. The skin of the epidermis has excellent curative effect. Its biggest feature is that it can promote the proliferation and differentiation of cells, thereby replacing aging and dead cells with new cells. EGF can also stop bleedin...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P21/02C07K1/16C07K1/14A61K38/18A61K8/64A61P17/02A61P3/10A61P27/02A61Q1/00
Inventor 孙民富
Owner 孙民富
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