Date: Wednesday, October 17th, 2018

Title: Second Attempt Plaque Assay for Soil Sample C

Rationale: The purpose of today’s lab is to make a plaque assay for a new soil sample in order to search for plaques.

Class Question: Is there a difference in bacteriophage presence or type in soil samples taken from live oaks vs those from red oaks?

Procedure:

1. An aseptic zone was set up
2. Plaque assays were retrieved and evaluated. They were found to be inconclusive and stored in the walk-in freezer.
3. Another 10 microliters of enriched lysate was added to a culture tube with .5 mL arthrobacter and left to sit for 15 minutes.
4. Agar was made for three plates with the following formula:
   1. 6 mL LB broth
   2. 7.5 mL 2x TA
   3. 67.5 microliters 1M CaCl2
5. 4.5 mL of the TA solution was added to each culture tube and the TA control plate.
6. The plates were left for 20 minutes to harden before being inverted and incubated for the next 5 days.

Observations: The plaque assay and the TA control were not contaminated and there were spots on the agar, implying the presence of phage; however, the spots were misshapen and the plate had not fully solidified. It’s possible that plaques were present and had become abnormal due to the liquidity of the plate.

Results: This experiment yielded a new plaque assay that can be evaluated at a later date for plaques.

Next Steps: The next step is to evaluate the plaque assay. If it yields positive results, the next step is to pick a plaque. If negative, a new sample may have to be adopted.