Assessing the Antimicrobial Potential of Bacteriocin-Producing Murine Fecal Bacterial Isolates

To count bacteriocin-producing bacterial cells in fecal samples, transfer diluted cells of the bacteriocin-producer to 4 milliliters of pre-warmed MRS soft agar. Mix the agar by vortexing, and pour the cells onto an MRS 1.5% agar plate. This layer is referred to as the first layer.

Transfer another 4 milliliters of cell-free 0.8% MRS soft agar onto the first layer to embed all the cells within the soft agar. This layer is meant to prevent cells from growing as colonies on the surface of the agar plates.

In the sterile hood, dry the plates by taking the lid off for five to 10 minutes before incubating the plates at 30 degrees Celsius for 20 to 24 hours for cell growth and colony formation.

To identify bacteriocin-producing colonies, mix 40 microliters of an overnight culture of an appropriate indicator strain with 4 milliliters of pre-warmed soft agar for each plate. Pour the mixture onto the plates. This layer is referred to as the third layer.

Dry and incubate the plates as before, for cell growth and colony formation. Bacteriocin-producing colonies have clear zones of inhibition around the colonies.