Question 1

How much nutrient agar is needed to prepare bacterial culture plates?

Accepted Answer

Add 23 grams of nutrient agar to a 2,000 mL beaker containing 1 liter of distilled water. Place the beaker on a hotplate set to high and stir until the agar dissolves completely. This quantity prepares enough nutrient agar medium for multiple Petri dishes used in microbial culture experiments.

Question 2

What is the proper procedure for sterilizing nutrient agar before plating?

Accepted Answer

After the agar dissolves, transfer the beaker to an autoclave and run it at 121°C for 15 minutes. Remove the beaker and allow it to cool to the touch for about 15 minutes without letting the agar solidify. This sterilization process eliminates contaminants and prepares the medium for safe bacterial and yeast culture.

Question 3

How should you prepare Petri dishes before inoculating them with bacteria?

Accepted Answer

Pour the cooled liquid nutrient agar into each Petri dish until it is about halfway full. Allow the agar to solidify, then seal the sides with Parafilm. Under a fume hood, use a marker to draw two intersecting lines on the bottom of each plate to divide the agar into four equal quadrants, and label with initials, bacterial species name, and date.

Question 4

Why is flame sterilization important when using an inoculating loop?

Accepted Answer

Flame sterilization eliminates microbial contamination from the inoculating loop between samples. Heat the looped end in a Bunsen burner flame for 20 seconds, then allow it to cool for about 15 seconds before use. This prevents cross-contamination between bacterial cultures and ensures pure, isolated colonies on your plates.

Question 5

What is the streak plating technique and how does it isolate bacterial colonies?

Accepted Answer

Streak plating involves spreading bacteria across quadrants in a specific pattern where each new streak crosses the previous one only once or twice. This dilutes the bacterial population progressively, allowing isolated single colonies to grow in later quadrants. The technique is essential for obtaining pure cultures and observing microbial colony and fungal diversity in laboratory settings.

Question 6

How should yeast cultures be plated differently from bacterial cultures?

Accepted Answer

For yeast plating, scoop S. cerevisiae from a vial onto the agar using an inoculating loop and evenly spread it across the entire plate, rather than using the streak plating method. Label the plate with the yeast strain name, date, and your name. Reseal with Parafilm and store upside down at 32°C until the activity.

Question 7

What storage conditions are required for bacterial and yeast cultures after plating?

Accepted Answer

After sealing plates with Parafilm, store bacterial cultures lid down in a 32°C incubator until the day of the activity. Yeast cultures should also be stored at 32°C but positioned upside down. Proper incubation temperature and orientation promote optimal microbial growth while preventing condensation from dripping onto colonies.

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