Microbiologists often use the technique of inoculation to grow cultures of micro-organisms. For organisms grown in petri dishes, a solid agar medium provides the nutrients necessary for the microbes to survive. Inoculation requires only a few basic laboratory techniques, but it is imperative that you carefully follow sterile techniques to prevent contamination of the petri dish or the microbial culture.
- Skill level:
Things you need
- Petri dish containing prepared agar
- Microbial culture
- Bunsen burner
- Inoculating loop
Move all the materials for the inoculation into a laminar flow hood if you have access to one.
Hook the Bunsen burner hose up to a gas nozzle. Turn on the gas and light the burner.
Flame the inoculating loop by holding the loop in the flame of the Bunsen burner for several seconds.
Obtain a sample of the microbial culture. If it is a broth culture, remove the lid from the culture tube--to prevent contamination, do not set the lid down. Pass the opening of the tube through the flame of the Bunsen burner for one second or less. Insert the inoculating loop into the broth and draw it out. Flame the opening of the tube again and replace the lid.
If the culture is on an agar plate or agar slant, remove the lid just far enough to insert the inoculating loop and scrape the loop gently over the surface of one microbial colony to obtain some of the material. Replace the lid.
Lift the lid of the petri dish that you will inoculate to about a 45 degree angle. Streak the inoculating loop containing the microbial culture across the surface of about one quarter of the agar. Replace the lid.
Use the quadrant streak method to isolate colonies of the microbe. Flame the inoculating loop, lift the lid of the petri dish, streak from one edge of the previous streak across another quarter of the plate, and replace the lid. Repeat this procedure twice.
Invert the petri dish and incubate it for 24 to 48 hours.
Tips and warnings
- Don't set the inoculating loop down or touch anything with it after flaming, or it may become contaminated.
- Use disposable plastic inoculating loops to eliminate the need for flaming--use a new loop whenever the procedure calls for flaming.
- Follow aseptic technique and laboratory protocols throughout the inoculation.
- Properly store or dispose of the microbial cultures when you are done with the experiment.
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