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Fermentation method and fermentation medium for recombinant human III type collagen engineering bacteria

A collagen and fermentation method technology, applied in the direction of animal/human protein, microorganism-based methods, fermentation, etc., can solve the problems of low collagen yield and insufficient large-scale production, and achieve the effect of ensuring growth vitality

Inactive Publication Date: 2017-03-22
河南多美康生物科技有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the yield of the target product, human collagen, is low enough for large-scale production

Method used

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  • Fermentation method and fermentation medium for recombinant human III type collagen engineering bacteria
  • Fermentation method and fermentation medium for recombinant human III type collagen engineering bacteria

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0048] The operation steps of the fermentation method of the recombinant human type III collagen engineering bacteria provided in this example are as follows.

[0049] 1 activated strain

[0050] 1.1 Take the cryopreserved recombinant human type III collagen engineering bacteria, after thawing, inoculate the recombinant human type III collagen engineering bacteria into YPD solid medium, and grow at 28°C for 48 hours until the diameter of a single colony is 3mm about.

[0051] Among them, the recombinant human type III collagen engineering bacteria used in this example is transformed with the recombinant expression vector pPICZaA yeast (Pichia Pastoris), wherein pPICZaA carries the human collagen gene (Col3-3) encoding human collagen, the restriction site at the 5' end of the gene is XhoI, and the restriction site at the 3' end is SacII.

[0052] Biological preservation information of recombinant human type III collagen engineering bacteria used in this example: Pichia Pasto...

Embodiment 2

[0069] The operation steps of the fermentation method of the recombinant human type III collagen engineering bacteria provided in this example are as follows.

[0070] 1 activated strain

[0071] The specific steps of activating the bacteria in this example are basically the same as those in Example 1, so as to obtain the seed solution of recombinant human type III collagen engineered bacteria.

[0072] 2 Fermentation culture

[0073] 2.1 Inoculate the obtained recombinant human type III collagen engineered bacteria seed solution at a volume ratio of 1:10 into a fermenter containing 40L of basic fermentation culture medium (Shanghai Bailun, model: BLBIO-100STA-UIP, total capacity 60L) . The fermenter can display dissolved oxygen, pH, temperature, stirring speed and other parameters, and is equipped with a peristaltic pump for automatic pH control and automatic feeding.

[0074] Wherein, the basic fermentation medium used in the present embodiment comprises by every liter: H...

Embodiment 3

[0083] The operation steps of the fermentation method of the recombinant human type III collagen engineering bacteria provided in this example are as follows.

[0084] 1 activated strain

[0085] The specific steps of activating the bacteria in this example are basically the same as those in Example 1, so as to obtain the seed solution of recombinant human type III collagen engineered bacteria.

[0086] 2 Fermentation culture

[0087] 2.1 Inoculate the obtained seed liquid of recombinant human type III collagen engineering bacteria into a fermenter containing 40L of basic fermentation culture medium at a volume ratio of 1:8 (Shanghai Bailun, model: BLBIO-100STA-UIP, total fermentation capacity 70L ). The fermenter can display dissolved oxygen, pH, temperature, stirring speed and other parameters, and is equipped with a peristaltic pump for automatic pH control and automatic feeding.

[0088] Wherein, the basic fermentation medium used in the present embodiment comprises by ...

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Abstract

The invention discloses a fermentation method and fermentation medium for recombinant human III type collagen engineering bacteria and belongs to the technical field of engineering bacteria fermentation. The fermentation method comprises the following steps that the recombinant human III type collagen engineering bacteria are inoculated to a basic fermentation culture solution to conduct fermental cultivation, and the basic fermentation culture solution comprises, by liter, 25-27 ml of H3PO4, 0.8-1.0 g of CaSO4, 17-19 g of K2SO4, 13-15.5 g of MgSO4.7H2O, 3.8-4.3 g of KOH, 37-43 g of glycerinum and 3.5-4.2 ml of a first microelement solution; and in the fermental cultivation process, supplementary materials containing glycerinum and a second microelement solution are added into the basic fermentation culture solution. By optimizing the basic fermentation culture solution and the component content of the supplementary materials, the protein expressing quantity of the recombinant human III type collagen engineering bacteria is greatly improved by the fermentation method, and the method can be applicable to industrial large scale production.

Description

technical field [0001] The invention relates to the technical field of engineering bacteria fermentation, in particular to a fermentation method and fermentation medium of recombinant human type III collagen engineering bacteria. Background technique [0002] At present, the existing Escherichia coli fermentation method used to produce human collagen (Human Collagen) is mainly produced in the form of inclusion bodies. Although this method can produce high yields, it will cause complications in terms of inclusion body refolding and subsequent purification. The cumbersome process limits the large-scale production of the target protein. Soluble expression systems are also currently available for the production of soluble human collagen, which, however, avoids the problem of inclusion bodies. However, the yield of the target product human collagen is very low, which is not enough for large-scale production. Contents of the invention [0003] The purpose of the present invent...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12P21/02C12N1/16C12R1/84
CPCC07K14/78C12N1/16
Inventor 贾芳苗梁波邓飞
Owner 河南多美康生物科技有限公司
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