International A Level Biology / Microbiology

Culturing microorganisms with aseptic technique

Edexcel International A Level Biology (YBI11), chapter 17: Microbiology, specification point 6.1. Free lesson.

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Aseptic technique lets you grow the microorganism you want while limiting contamination and keeping the culture safe. You need to know how to prepare and inoculate plates, and why each safety step is used.

How to prepare sterile media and plates

A nutrient medium provides the materials microbes need to grow. Nutrient broth is liquid, while agar sets it into a firm gel for plates. The medium and equipment must start sterile, so unwanted microbes do not grow alongside the intended organism. Nutrient broth contains glucose, amino acids and minerals in water. These provide carbon and nitrogen. Agar is a seaweed jelly added to broth to make a solid surface.

How to make a streak plate

A streak plate spreads microbes across the agar in stages. Each new section receives fewer cells, so separate colonies can form in the final section. A colony is a mound of cells that grew from one cell or a small clump. Use a wire inoculating loop to transfer an unmeasured loopful of culture. Flame the loop until it glows red to kill microbes on it, then let it cool. If it is still red hot, it will kill the bacteria you are trying to transfer.

How a spread plate differs

A spread plate distributes the sample evenly over the whole agar surface. It can produce an even lawn of growth, or separate colonies that can be counted if the sample is dilute. A streak plate uses a wire loop and an unmeasured sample. It is used to obtain separate colonies and a pure culture of one species. A spread plate uses a glass or plastic spreader and a measured volume, for example 0.1 cm³, added with a sterile pipette. A glass spreader is sterilised by dipping it in ethanol and then burning off the ethanol.

What each aseptic step protects

Aseptic technique means working in a way that prevents contamination by unwanted microorganisms. Some steps protect the culture; others protect you by preventing live microbes from leaving the laboratory. Autoclave media and glassware before use so only the intended microorganism grows. Autoclave used cultures before disposal so no live microbes leave the lab. Flame the loop before use to kill contaminants, and after use to kill the culture left on it. Work beside a Bunsen burner. Rising warm air carries microbes away, and a slightly lifted lid lets fewer airborne microbes land on the…

How to explain a precaution

An exam answer should say what is protected and how the precaution works. Saying only “to avoid contamination” does not explain what could be contaminated or what would happen. For example, flaming the neck of a culture tube kills microbes on its rim, so they do not enter the culture and grow alongside the intended organism. To protect a streak plate, flame the loop until red hot and let it cool. This kills microbes on the loop, so only the intended organism is transferred. Lift the lid only slightly. This reduces the number of airborne microbes that can land on the agar.

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