Results:

No contamination, however, the “KEA 11/5 PA 10^-2” plate looked strange since it folded up when solidifying.

Rationale:

The titer will be calculated from the “KEA 11/5 PA 10^-1” plate. From this, the amount of lysate needed to webbed the plate will be calculated. Three plaque assays will be run with slightly varying amounts to increase the likelihood of receiving a webbed plate.

Procedure:

1. Counted the total number of plaques on “KEA 11/5 PA 10^-1” plate.
2. Calculated the average diameter of plaques, the titer, and the amount of lysate needed to web the plate.
3. Once an aseptic zone was established, 97 µL, 125 µL, and 150 µL of “KEA 11/5 FS lysate 10⁰” were added to correlated test tubes which already contained 0.5 mL of Arthrobacter.
4. 8 mL of LB Broth, 90 µL of CaCl₂, and 10 mL of 2X TA were combined into a conical vial.
5. Transferred and mixed 4.5 mL of the Top Agar (TA) mixture from the conical vial into each test tube.
6. Each test tube was poured onto their correlating plate, and the remaining 4.5 mL of the TA mixture was poured onto a plate labeled “KEA 11/7 Control.”
7. These plates were placed in the incubator at room temperature.

Observations:

• 132 plaques were counted from the “KEA 11/5 PA 10^-1” plate. The following photos show the plates from previously.

• The following measurements were recorded from 10 random plaques to approximate the average diameter: 0.3, 0.3, 0.5, 0.6, 0.5, 0.6, 0.2, 0.2, 0.3, and 0.3 mm. The average diameter calculated was 0.38 mm.
• The approximate plaque diameter from the “KEA 11/5 PA 10^-1” plate were slightly less than half the size of the approximate plaque diameter from the “KEA 10/29 10⁰PA” plate.

Calculations:

• After the plaque assays had been performed, it was noticed that the webbed calculations were done using the plaque diameter instead of the radius. The following shows the correct calculations.

• The following calculations were performed to determine enough LB Broth, 2X TA, and CaCl₂needed for 4 plates.
  

  Original Recipe	X4
  2 mL LB Broth	8 mL LB Broth
  2.5 mL 2X TA	10 mL 2X TA
  22.5 μL CaCl2	90 μL CaCl2

  Next Steps:

  In open lab, plaque assays will be performed with the correct amount of lysate.