Title: Plaque Picking and Metadata Results

Date: 10 October 2018

Rationale: The plaque assay from the previous lab appears to have potential plaques. The plaques will be picked and 2 more plaque assays will be done to attempt to isolate a phage.

Procedure: Aseptic zone created by washing the lab bench with CiDecon and Ethanol, and an ethanol lamp was lit for use in the workspace.

• 2 different plaques were picked with a micropipet tip and placed into individual vials of 100 microliter Phage Buffer (creating 2 different lysates).

The following recipe was used to make 2 plaque assays:

• 5 mL 2X Top Agar
• 4 mL LB Broth
• 45 microliters CaCl2

~4. 5 mL transferred into tube containing Arthrobacter + 10 microliters of lysate. Both plates were cooled for 15 minutes and placed into the incubator for 48 hours.

Metadata Results:

• % Water-  the final mass of the dirt was 10.858 g. Therefore the calculation (14.381-10.858)/14.381 = .2449 means that the soil was ~24.5% water.
• % Sand, Silt, Clay – 2.5 mL of separated sediment was visible in the falcon tube, giving the following results:
  • 1 mL Sand | 1mL/3mL = 33.33% sand
  • 1.5 mL Silt | 1.5mL/3mL = 50% silt
  • 0.5 mL Clay | 0.5mL/3mL = 16.67% clay

Conclusions: 2 different plaques were picked on the resulting plate to maximize the chances of finding a phage that can be passaged in a later lab session. The % water test yielded a seemingly high proportion of water, which can be explained by the recent rainfall that likely saturated the soil. The first picked potential plaque appeared to clear out most of the Arthro on the plate while the second picked potential plaque appeared as a small, black circle on the other side of the plate. Attached is a picture of the plate from October 8th.