Microbiology Lab Experiment Changes
Experiment #: 6-3
Title: Bacteriophage Plaque Assay
Live Organisms: E. coli B, Bacteriophage T2 or T4
Changes: Procedure (Work in groups)
1. Soft agar deeps have been melted and are being held
at 45 - 50°C in water bath.
2. Use only 4 nutrient broth tubes not 7 as in manual. You
are starting with 10-4 phage dilution. Use only 5 TSA
plates.
3. Keep soft agar tubes in little water baths. Perform
transfers in the water baths. It is a good idea to put the
broth tubes in the water baths also. This way everything is
the same temperature.
4. Follow procedure in lab manual except transfer 1.0 mL
of each phage dilution to the soft agar deeps not 0.1mL as
in lab manual and omit the second row of tubes. You will
be putting the phage dilution and E. coli directly into the
soft agar tubes.
Take Home Lesson: Read lab manual for review. Define: plaque, PFU, TNTC,
and TFTC. As with the previous serial dilution, the number of plaques must fall
between 30 and 300. Calculate the number of pfu’s per mL of stock phage culture
by multiplying the number of pfu’s on a plate times the dilution factor of the phage
placed on that plate. What type of phage are we using? What life cycle does it
use? Explain the steps in this life cycle.
