Culturing Bacteria - Lab #2: Enrichment of Microbes Using a Milk Medium

Culturing Bacteria
Activity #2

Enrichment of Microbes Using a Milk Medium

Be sure to read the General Safety Guidelines and the Microbiology Safety Guidelines before beginning this lab activity.

Milk is an excellent medium for growing a variety of microorganisms (think sour milk!). The following procedure is designed to allow for the isolation of one of the organisms that flourishes in the milk medium.
Culturing Bacteria Menu
Introduction
Activity #1: Bacterial Enrichment
Using a Carrot Medium
Activity #2: Enrichment of Microbes
Using a Milk Medium
Appendix
General Safety Guidelines
Microbiology Safety Guidelines

Materials and Apparatus

Fresh milk
Curdled sour milk
Transfer loop
5 test tubes
5 sterile petri dishes
Calcium carbonate (CaCO₃)
Agar

Procedure

Transfer a loopful of curd from sour milk to a test tube containing sterile water. Label this tube #1.

Perform a series of dilutions by transferring a loopful of liquid from tube #1 to a second tube containing sterile water. Label this tube #2.

Repeat step #3 for three more tubes, #3,#4, and #5.

Prepare 5 sterile petri dishes with sterile milk isolation medium (see separate preparation formula).

Place a loopful of inoculum from each of the tubes (steps #2-#4) onto HALF of each plate (use standard streaking technique to apply the inoculum).

Repeat the inoculation of the same 5 plates but apply the inoculum BELOW THE SURFACE of the agar. To do this, use a sterile inoculation loop for each test tube of diluted microbes and transfer the inoculum to the SECOND HALF of the plate by pushing the loop of the transfer device BELOW the surface of the agar. Make only one indentation per plate.

Incubate the 5 plates at 30慢 overnight.

Make slide preparations and Gram stain the individual colonies.

Acid production by the microbes is indicated by the decomposition of CaCO₃ that was added to the agar. This decomposition produces haloes around the colonies.