I recently experimented with T4 bacteriophage in a microbiology lab. My findings were as follows:
This lab involved two experiments, the titration and isolation of a lytic phage and a phage typing experiment.
In the experiment with the titration and isolation of a lytic phage, it was predicted that as the lytic phage became more diluted, it would produce less plaques on the Escherichia coli lawns, and that at least one plate would produce a countable number of plaques. This experiment was performed by creating a serial dilution of the T4 phage. This phage was diluted to 10^-8 dilution by initially combining 0.1 mL of T4 phage with 0.9 mL of TM buffer, then taking .1 mL of this solution and transferring it into another tube with0 .9 mL of TM buffer. This process was repeated until the 10^-8 dilution was achieved. Each of these solutions then had 0.1 mL transferred onto its own individual Luria agar bottom plate. Each of these plates also had 0.1 mL of E. coli B/r poured onto them. These solutions were swirled around the plate to make sure all of the surface area was covered. After these bacteria and phages were allowed some time to grow on the plates, the plates were inspected for plaques, which are small holes in the bacteria lawn caused by phage lysing. Plates with over 300 plaques were considered too numerous to count, and plates with less than 30 plaques were considered too few to count.
Dilution levels 10^-1 through 10^-5 were too numerous to count, and dilution level 10^-8 was too few to count. Dilution level 10^-6 had 205 plaques, and dilution level 10^-7 had 73 plaques. The plaque forming units/milliliter were then calculated using the equation average of PFU/ amount of phage added to the plate * dilution factor. The plate at dilution factor -6 was found to have 2050000000 PFU/mL, and the plate at dilution factor -7 was found to have 7300000000 PFU/mL.
The expected results of this experiment were confirmed, two plates with a countable number of plaques were cultivated.
In the phage typing experiment, it was expected that the two E. coli strains would be infected by T4 phage, as E. coli have binding sites on their cell surface that T4 bacteriophage can bind to and infect the cell from. A LB plate was divided into four quadrants, and each was swabbed with its own individual bacteria (E. coli B/r, E. coli K12, Salmonella arizonae, and P. vulgaris). Each of these tiny lawns was then inoculated with T4 phage from the original plate, using a toothpick to stab the burst zone on the original plate and transfer it to the new plate. A new toothpick was used for each lawn in an effort to prevent cross contamination. E. coli K12 and E. coli B/r both developed burst zones around the area of inoculation, and Salmonella arizonae, and P. vulgaris did not, showing that the T4 phage only affected E. coli K12 and E. coli B/r. The expected results were confirmed, as both strains of E. coli were infected with T4 bacteriophage.