Fermentation medium and fermentation method for preparing recombinant human interferon alpha2b

Abstract

The invention discloses a fermentation medium and a fermentation method for preparing recombinant human interferon alpha2b, including a fermentation medium formula which is suitable for constitutive expression recombinant human interferon alpha2b engineering bacteria and a fermentation method thereof; the fermentation medium formula comprises the following components in parts by weight: 25-35 parts of tryptone, 15-25 pats of yeast extract, 4-8 parts of disodium hydrogen phosphate, 2-4 parts of mono-potassium phosphate, 0.4-0.6 parts of ammonium chloride, 0.4-0.6 parts of sodium chloride, 0.2-0.4 parts of magnesium sulfate, 0.05-0.15 parts of calcium chloride and 10-20 parts of glycerol; the glycerol is used as the only carbon source during a culture process of the fermentation method, and the expression rate of target protein is regulated by using proper phosphate concentration, therefore the excellent growing status of the engineering bacteria is ensured, and the high-density culture and high-efficiency expression are achieved.

Description

technical field [0001] The invention relates to the field of medicine, in particular to a fermentation medium and a fermentation method for preparing recombinant human interferon α2b. Background technique [0002] With the development of genetic engineering technology, the fermentation process of engineering bacteria has been widely used. The purpose of bioengineering bacteria fermentation is to obtain a large number of exogenous gene products and reduce the pollution of host cells and external factors as much as possible. Therefore, high density The cultivation of high yield and high expression has become the goal and direction of the fermentation industry. In terms of high-density expression, Escherichia coli is the most representative and widely used host bacteria. There are two ways to express recombinant proteins in E. coli, one is constitutive expression and the other is inducible expression. In the high-density fermentation culture of protein engineering bacteria, mo...

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Problems solved by technology

In terms of high-density expression, Escherichia coli is the most representative and widely used host bacteria. There are two ways to express recombinant proteins in E. coli, one is constitutive expression and the other is inducible expression. In the high-density fermentation culture of protein engineering bacteria, most of them are inducible expression strains. The advantage is that through temperature induction or chemical induction, the engineering bacteria can grow rapidly in a short period of time, achieve a high specific growth rate, and achieve maximum expression at the same time. The disadvantage is that during the rapid growth of bacteria, the metabolites of bacteria will also be produced rapidly, such as the rapid accumulation of harmful substances such as organic acids and carbon dioxide, which will inhibit the growth of bacteria and the expression of products, and the production of the largest by-product, acetic acid, It is generally believed that under the condition of low specific growth rate, the energy produced by oxidative metabolism of bacteria is sufficient to meet the needs of synthesis and dissimilation, and will not produce acetic acid, while at high specific growth rate, Escherichia coli cannot provide enough energy by oxidative metabolism alone. The energy must be stored in ATP and NADH through the acetic acid production pathway, so that excessive acetic acid will be produced, and the increase in acid will inhibit bacterial reproduction and product expression, and will also bring a great burden to subsequent purification

[0003] The constitutive expression method of recombinant protein in Escherichia coli is a slow expression method without induction, and it continues to grow and express during the growth of the bacteria. Because there is no induction of rapid growth during the growth cycle, the growth rate is uniform and the metabolites are Low acetic acid content can achieve ideal product expression rate, but it is difficult to achieve high density culture

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