7 November 2018 ✷ Pick a Plaque Re-do

Rationale: Members of group 6 adopted phage from Soil 4 from Melissa (collected 10/5/18) in order to help amplify the phage sample. A plaque assay was run (pick a plaque) to amplify present phage. The first plaque assay showed heavy contamination so it will be redone in order to have a plate ready for flooding.

Procedure

• The table was cleaned with Cidecon and 70% ethanol and an ethanol lamp was lit.
• The lysates from Monday’s lab were still available and there was enough arthrobacter for every group member to make a plate. One large vial was mixed with the following volumes and concentrations:

• component	volume	final concentration
  LB Broth	8 mL	–
  2X Top Agar	10 mL	1X
  1M Calcium Chloride	90 µL	4.5 µM

• 30 µL of the lysate was allowd to infect the arthrobacter for 15 minutes before being combined with 5 mL of the above mixture and poured into a plate, allowed to harden, and incubated until Monday.

Observations, results, data

The contamination seen in the plates likely came from the top agar or LB broth because several of the other groups that made plates experienced the same contamination.

There were very few plaques in the plaque assay run on Monday, so the volume of lysate was increased in the plaque assay run on Wednesday.

Interpretations, conclusion, next steps

If this plaque assay is run without contamination in the control plate, it will be amplified by flooding the plate with phage buffer in order to increase the concentration of phage in the sample.