OVERVIEW
Lambda-agar is nothing but a lambda broth which is solidified by agar. Agar is a solidifying agent. It melts at 85°C but melton agar stays liquid at a temperature > 40°C (this property is called hysteresis, different temperatures for melting and solidifying). This allows us to add heat-labile medium supplements such as antibiotics and vitamins and prepare solid medium.
Lambda-agar solid medium is prepared by dissolving 15 g agar / liter of Lambda broth (Elbing & Brent, 2019). Whenever needed, solid medium is melted and plates are poured.
Here we describe a procedure to prepare 500 ml Lambda-agar solid medium that is sufficient to prepare ≈ 15 plates (≈ 30 ml medium/10 cm petri-plate).
REQUIREMENTS
Reagents and Solutions
Lambda broth
Agar
Equipment and disposables
Erlenmeyer flask
Hot plate
Composition
1x Lambda broth
1.5% Agar
Objective:
Preparation of 500 ml of Lambda-agar medium
PREPARATION
Step 1: Weigh out 7.5 g agar in a 1-liter Erlenmeyer flask/autoclavable bottle. Add 500 ml Lambda broth. Mix it by swirling the flask.
Step 2: Close the lid of the bottle loosely/insert a cotton plug and cover the mouth of the Erlenmeyer flask with aluminum foil.
Step 3: Sterilize the solution by autoclaving for 20 minutes at 15 lb/in2 (1.05 kg/cm2) on liquid cycle.
Important notes:
Lambda-agar medium (solid at room temperature) can be stored at room temperature for a few weeks but for a longer time, we recommend you to store it in the cold room.
Depending on your convenience, you can prepare the lambda-agar plate any time by melting the medium.
After autoclaving, the medium must be handled aseptically to maintain sterility.
Step 4: Allow the medium to cool below 50°C. If required, add supplements such as antibiotics. Mix by swirling the flask gently.
Step 5: Pour ≈ 30 ml medium per 10 cm plate. Let the medium solidify. Keep the plate open for 2 h at room temperature to dry the plate.
Tip:
Remove the trapped air by touching with the needle/micropipette tip.
Precaution:
Don’t cover the plate with a lid. If the plate is covered with the lid, all the vapour will condense on the lid and form water droplets that eventually fall back to the solidified medium.
Storage:
Cover the plates with their lid and store at 4°C in an inverted position.
REFERENCES
- Elbing & Brent, 2019. Recipes and Tools for Culture of Escherichia coli. Curr Protoc Mol Biol. 125(1), e83. PMID-30412361; Full-Text Links: wiley, PMC
