Recombinant escherichia coli shake flask culture medium and method for increasing expression quantity of target protein

Abstract

The invention provides a recombinant escherichia coli shake flask culture medium, which is prepared from the following components: soy peptone, yeast powder, ammonium sulfate, potassium dihydrogen phosphate, disodium hydrogen phosphate, ginkgo biloba seed polysaccharide, magnesium potassium sulfate and sodium naphthalene acetate. The invention also optimizes the culture conditions and the culture way of a strain in shake flask culture, and establishes a shake flask expression method which can enable thallus to grow with high density and the target protein to be expressed efficiently at the same time. In the premise of increasing the amount of the thallus per unit volume, the total protein content per unit volume is increased, so that the yield of the target product is increased, and the specific productivity of the product (product yield per unit volume and per unit time) is finally increased; the recombinant escherichia coli shake flask culture medium not only can reduce the culture volume and strengthen the downstream separation and extraction, but also can shorten the production cycle and reduce the equipment investment so as to reduce the production cost, so that the competitiveness of the recombinant escherichia coli shake flask culture medium in the market is greatly enhanced.

Description

technical field

[0001] The invention relates to the field of biotechnology, in particular to a recombinant Escherichia coli shake flask culture medium and a method for increasing the expression level of a target protein. Background technique

[0002] The prokaryotic expression system has the characteristics of short expression period, high expression level, simple operation, easy cultivation, and low cost. Therefore, it is usually used as an expression system for exogenous genes in DNA recombination technology, and the E. coli expression system is currently the most mature technology. Prokaryotic expression system.

[0003] For the expression of recombinant bacteria for the purpose of producing foreign proteins, it is hoped to obtain as much target protein as possible during the expression process, improve the volume production efficiency of the product, and increase the yield of the final product. In the process of expressing foreign proteins, recombinant engineering bacte...

Examples

Embodiment 1

[0025] A recombinant Escherichia coli shake flask culture medium, comprising the following preparation method: weighing the following components: 20g of soybean peptone, 10g of yeast powder, 2.5g of ammonium sulfate, 2.3g of potassium dihydrogen phosphate, 11.84g of disodium hydrogen phosphate, and 3g of ginkgo polysaccharide , Potassium magnesium sulfate 1g, sodium naphthalene acetate 12g, add purified water to dissolve and make up to 900mL, then add 0.1mL antifoaming agent, after preparation, sterilize at 121°C for 30 minutes to obtain.

[0026] The method for improving the expression of recombinant protein by applying the described recombinant Escherichia coli shake flask culture medium comprises the following steps:

[0027] a. Take 50 mL of LB culture solution, add 50 uL of 35 mg / mL kanamycin sulfate solution, inoculate the bacteria containing the recombinant protein into the LB culture solution, and culture overnight at 37°C and 50 rpm after inoculation;

[0028] b. Take...

Embodiment 2

[0032] A recombinant Escherichia coli shake flask culture medium is characterized in that it comprises the following preparation method: weighing the following ingredients: 15g of soybean peptone, 15g of yeast powder, 2g of ammonium sulfate, 3g of potassium dihydrogen phosphate, 10g of disodium hydrogen phosphate, ginkgo polysaccharide 2g, 1g of potassium magnesium sulfate, 12g of sodium naphthalene acetate, add water to dissolve and make up to 900mL, then add 0.3mL of antifoaming agent, after preparation, sterilize at 121°C for 45 minutes to obtain the product.

[0033] The method for improving the expression of recombinant protein by applying the described recombinant Escherichia coli shake flask culture medium comprises the following steps:

[0034] a. Take 50 mL of LB culture solution, add 50 uL of 30 mg / mL kanamycin sulfate solution, inoculate the bacteria containing the recombinant protein into the LB culture solution, and culture overnight at 37°C and 70 rpm after inocul...

Embodiment 3

[0039]A recombinant Escherichia coli shake flask culture medium, comprising the following preparation method: weighing the following ingredients: 30g of soybean peptone, 8g of yeast powder, 4g of ammonium sulfate, 1g of potassium dihydrogen phosphate, 15g of disodium hydrogen phosphate, 3g of ginkgo polysaccharide, potassium sulfate Add 2g of magnesium and 10g of sodium naphthaleneacetate, dissolve in water and make up to 900mL, then add 0.1-0.3mL of antifoaming agent, and sterilize at 121°C for 45 minutes after preparation.

[0040] The method for improving the expression of recombinant protein by applying the described recombinant Escherichia coli shake flask culture medium comprises the following steps:

[0041] a. Take 50 mL of LB culture solution, add 50 μL of 50 mg / mL kanamycin sulfate solution, inoculate the bacteria containing the recombinant protein into the LB culture solution, and culture overnight at 37°C and 70 rpm after inoculation;

[0042] b. Take 100mL of 20% ...

technical field

[0001] The invention relates to the field of biotechnology, in particular to a recombinant Escherichia coli shake flask culture medium and a method for increasing the expression level of a target protein. Background technique

[0002] The prokaryotic expression system has the characteristics of short expression period, high expression level, simple operation, easy cultivation, and low cost. Therefore, it is usually used as an expression system for exogenous genes in DNA recombination technology, and the E. coli expression system is currently the most mature technology. Prokaryotic expression system.

[0003] For the expression of recombinant bacteria for the purpose of producing foreign proteins, it is hoped to obtain as much target protein as possible during the expression process, improve the volume production efficiency of the product, and increase the yield of the final product. In the process of expressing foreign proteins, recombinant engineering bacte...

Examples

Embodiment 1

[0025] A recombinant Escherichia coli shake flask culture medium, comprising the following preparation method: weighing the following components: 20g of soybean peptone, 10g of yeast powder, 2.5g of ammonium sulfate, 2.3g of potassium dihydrogen phosphate, 11.84g of disodium hydrogen phosphate, and 3g of ginkgo polysaccharide , Potassium magnesium sulfate 1g, sodium naphthalene acetate 12g, add purified water to dissolve and make up to 900mL, then add 0.1mL antifoaming agent, after preparation, sterilize at 121°C for 30 minutes to obtain.

[0026] The method for improving the expression of recombinant protein by applying the described recombinant Escherichia coli shake flask culture medium comprises the following steps:

[0027] a. Take 50 mL of LB culture solution, add 50 uL of 35 mg / mL kanamycin sulfate solution, inoculate the bacteria containing the recombinant protein into the LB culture solution, and culture overnight at 37°C and 50 rpm after inoculation;

[0028] b. Take...

Embodiment 2

[0032] A recombinant Escherichia coli shake flask culture medium is characterized in that it comprises the following preparation method: weighing the following ingredients: 15g of soybean peptone, 15g of yeast powder, 2g of ammonium sulfate, 3g of potassium dihydrogen phosphate, 10g of disodium hydrogen phosphate, ginkgo polysaccharide 2g, 1g of potassium magnesium sulfate, 12g of sodium naphthalene acetate, add water to dissolve and make up to 900mL, then add 0.3mL of antifoaming agent, after preparation, sterilize at 121°C for 45 minutes to obtain the product.

[0033] The method for improving the expression of recombinant protein by applying the described recombinant Escherichia coli shake flask culture medium comprises the following steps:

[0034] a. Take 50 mL of LB culture solution, add 50 uL of 30 mg / mL kanamycin sulfate solution, inoculate the bacteria containing the recombinant protein into the LB culture solution, and culture overnight at 37°C and 70 rpm after inocul...

Embodiment 3

[0039]A recombinant Escherichia coli shake flask culture medium, comprising the following preparation method: weighing the following ingredients: 30g of soybean peptone, 8g of yeast powder, 4g of ammonium sulfate, 1g of potassium dihydrogen phosphate, 15g of disodium hydrogen phosphate, 3g of ginkgo polysaccharide, potassium sulfate Add 2g of magnesium and 10g of sodium naphthaleneacetate, dissolve in water and make up to 900mL, then add 0.1-0.3mL of antifoaming agent, and sterilize at 121°C for 45 minutes after preparation.

[0040] The method for improving the expression of recombinant protein by applying the described recombinant Escherichia coli shake flask culture medium comprises the following steps:

[0041] a. Take 50 mL of LB culture solution, add 50 μL of 50 mg / mL kanamycin sulfate solution, inoculate the bacteria containing the recombinant protein into the LB culture solution, and culture overnight at 37°C and 70 rpm after inoculation;

[0042] b. Take 100mL of 20% ...

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