E. coli is typically grown in liquid culture by inoculating a sterile medium with a small number of cells. Here's a more detailed breakdown:
The Process of Growing E. coli in a Lab
Growing E. coli in a laboratory setting involves several key steps to ensure successful and reproducible results. This process is fundamental in various biological research fields.
1. Preparing the Growth Medium
The choice of growth medium is crucial. Commonly used media include:
- Lysogeny Broth (LB): A rich medium that supports rapid E. coli growth.
- Terrific Broth (TB): Another rich medium, often preferred for higher cell densities.
- Minimal Media (e.g., M9): Allows control over nutrient availability and specific metabolic studies.
The medium is prepared according to established protocols and then sterilized, typically by autoclaving, to eliminate any contaminating microorganisms.
2. Inoculation
A small number of E. coli cells from a stock culture (e.g., frozen stock or a colony from an agar plate) are introduced into the sterile growth medium. This process is called inoculation. Good aseptic technique is essential to prevent contamination.
3. Incubation
The inoculated medium is incubated at a controlled temperature, usually 37°C, which is optimal for E. coli growth. The culture is often placed on a shaker to ensure adequate aeration, promoting even growth.
4. Monitoring Growth
E. coli growth can be monitored by measuring the optical density (OD) of the culture using a spectrophotometer. OD is typically measured at 600 nm (OD600). As the cell density increases, the OD600 value also increases. This allows researchers to track the growth curve, which typically follows these phases:
- Lag Phase: Initial period where cells adapt to the new environment.
- Exponential (Log) Phase: Period of rapid cell division, with the population doubling every 20-30 minutes in rich medium.
- Stationary Phase: Growth slows due to nutrient depletion and accumulation of waste products.
- Death Phase: Cell death exceeds cell division.
Example Protocol: Growing E. coli in LB Broth
| Step | Description | Details |
|---|---|---|
| 1 | Prepare LB Broth | Dissolve LB powder in distilled water according to the manufacturer's instructions. |
| 2 | Sterilize the LB Broth | Autoclave at 121°C for 15-20 minutes. Let cool before use. |
| 3 | Inoculate the LB Broth | Using a sterile loop, transfer a single colony of E. coli from an agar plate into the sterile LB broth. |
| 4 | Incubate the Culture | Place the inoculated LB broth in a shaking incubator at 37°C and 200 rpm. |
| 5 | Monitor Growth | Measure the OD600 of the culture at regular intervals (e.g., every hour) to track growth. |
Alternative Growth Methods
While liquid culture is most common, E. coli can also be grown on solid media like agar plates. This allows for the isolation of single colonies, each originating from a single cell, which is useful for obtaining pure cultures. The general procedure involves streaking E. coli onto an agar plate and incubating it at 37°C until colonies form.
