When cultivating Escherichia coli in a fermentation tank, specific fermentation media are usually used, which are optimized according to the growth needs and production objectives of E. coli. The following are some commonly used fermentation media and their formulas: Common fermentation media for Escherichia coli mainly include the following:

1. LB medium (Lysogeny Broth)

LB medium is a commonly used E. coli culture medium, suitable for the growth, maintenance, and large-scale cultivation of E. coli.

Main ingredients:

·Tryptone: 10 g/L

·Yeast Extract: 5 g/L

·Sodium chloride (NaCl): 10 g/L

Features:

·Rich in nutrients, suitable for rapid growth of Escherichia coli.

·Agar can be added to make solid culture medium.

2. TB medium (Terrific Broth)

TB medium contains more nutrients than LB medium and is suitable for high-density fermentation.

Main ingredients:

·Tryptone: 12 g/L

·Yeast Extract: 24 g/L

·Glycerol: 4 mL/L

·Potassium dihydrogen phosphate (KH2PO4): 12.54 g/L

·Potassium dihydrogen phosphate (K2H2PO4): 8.56 g/L

·Ammonium chloride (NH4Cl): 3.4 g/L

Features:

·Provide higher levels of nutrients, suitable for high-density cell culture.

·Commonly used for protein expression and large-scale production.

3. SOB medium:

Suitable for protein expression in Escherichia coli, capable of increasing protein production, and also used for preparing SOC culture medium.

Main ingredients:

·2% tryptone

·0.5% yeast extract

·10 mM NaCl

·2.5 mM KCl

·10 mM MgCl2

·10 mM MgSO4

·pH7.5

Features:

·The ingredients are more complex, the nutrients are more abundant, and the nitrogen content is high, making it suitable for molecular biology experiments.

4. SOC culture medium:
Mainly used for the preparation and transformation of Escherichia coli competent cells, it can improve transformation efficiency and is also suitable for protein expression.

Main ingredients:

·2% tryptone

·0.5% yeast extract

·10 mM NaCl

·2.5 mM KCl

·10 mM MgCl2

·10 mM MgSO4

·20 mM D-glucose

·pH7.5

Features:

·Adding glucose to SOB medium provides a direct carbon source, which is beneficial for the growth of Escherichia coli.

5. M9 medium

M9 medium is a basic medium commonly used for specific experimental needs, such as the cultivation of nutrient deficient strains.

Main ingredients:

·葡萄糖 （Glucose）：4 g/L

·Ammonium chloride (NH4Cl): 1 g/L

·Sodium chloride (NaCl): 0.5 g/L

·Potassium dihydrogen phosphate (KH2PO4): 3g/L

·Sodium dihydrogen phosphate (Na2H2PO4): 6.78 g/L

·Magnesium sulfate (MgSO4): 0.241 g/L

·Calcium chloride (CaCl2): 0.011 g/L

Features:

·The nutritional composition is relatively simple and commonly used in basic research or experiments that require strict control of nutritional conditions.

6. 2xYT culture medium

2xYT medium is a variant of LB medium that contains higher nutritional content and is suitable for high-density cultivation.

Main ingredients:

·Tryptone: 16 g/L

·Yeast Extract: 10 g/L

·Sodium chloride (NaCl): 5 g/L

Features:

·Nutritious and suitable for high-density cultivation of Escherichia coli.

Supplements:

During the fermentation process of Escherichia coli, the following supplements may also need to be added:

·Antibiotics: used to select resistant strains.

·Inducing agents, such as IPTG, are used to induce the expression of recombinant proteins.

·Trace element and vitamin solutions: used to support cell growth.

During the fermentation process, the formula and conditions of the culture medium (such as pH, temperature, dissolved oxygen, etc.) need to be adjusted according to specific experimental purposes and production requirements. For example, if high-density fermentation is required, the amount of magnesium sulfate needs to be increased. Adding suitable yeast extract powder can increase the fermentation period, and adding ammonium sulfate can increase bacterial concentration. The fermentation process should be carried out under strict sterile conditions, and various parameters of the fermentation process should be monitored to ensure that the fermentation achieves the expected results.

The development process of Escherichia coli fermentation technology usually includes the following key steps:

1. Construction of bacterial strains:

Firstly, it is necessary to select an excellent strain, including genetic engineering route design, strain modification and screening, and exploration of shaking bottle conditions.

2. Optimization of culture medium:

The culture medium is an important component of the fermentation process of Escherichia coli. It is necessary to adjust the formula of the culture medium according to different strains and product requirements, and have a better understanding of the composition of the culture medium. DOE screening can be carried out, and the main components include the proportion of carbon sources, nitrogen sources, inorganic salts, and growth factors.

3. Process parameter optimization and control:

Temperature: The optimal temperature for the growth of Escherichia coli is usually 37 ℃, but the temperature range can be adjusted appropriately according to the fermentation purpose and strain characteristics.

PH value: The optimal pH value for the growth of Escherichia coli is 7 ± 0.2. During the fermentation process, it is necessary to maintain the pH stability of the fermentation broth through an acid-base feeding strategy.

Dissolved oxygen (DO): The critical dissolved oxygen concentration range for Escherichia coli is 20% to 30%. Insufficient dissolved oxygen can lead to the accumulation of acetic acid, affecting the growth of bacterial cells; Excessive dissolved oxygen may lead to oxygen poisoning. Therefore, it is necessary to control the dissolved oxygen level by adjusting parameters such as stirring speed and ventilation rate.

Feeding strategy: Select feeding batch fermentation to achieve high-density fermentation of Escherichia coli and obtain more biomass. During the feeding process, it is necessary to monitor the nutrient consumption in the fermentation broth in real time, develop the feeding process, determine the type, time, and amount of feeding.

During the development process of fermentation technology, software such as Minitab or JMP can be used to design DOE experimental methods to accelerate the experimental process and find better and more robust process parameters.