Title: Soil Collection and Washing

Date: 3 October 2018

Rationale: The previous dilutions all failed, therefore the conclusion was made that there was not a phage present in the previous sample. A new soil was collected to restart the phage discovery process.

Procedure: New soil was collected from a tree on the North side of Teal Residential hall. The tree was closer to the building, on the other side of the sidewalk. After soil was collected, it was washed and enriched.

Aseptic zone created by washing lab bench with CiDecon and Ethanol, and an Ethanol heat lamp was lit.

• 2 mL of fresh soil sample added to 15 mL conical vial
• LB Broth added to ~12 mL mark
• Vortex-ed for 15 minutes
• Centrifuged for 15 minutes at 3000 G
• Supernatant decanted into top-filter and vacuum filtered to yield ~9.5 mL direct lysate
• 0.5 mL Arthrobacter added to lysate to enrich
• Placed into shake incubator to complete enrichment process

Conclusions: The selected tree appeared slightly wilted with various dead branches, perhaps from a disease caused by a bacteriophage. The tree fit the criteria for the research question as it was likely planted by humans, not naturally grown like the large Oaks across campus. The metadata for the soil will be taken in the following lab session and the lysate tested for phage presence.