24 October 2018 ✷ Soil 6 Enrichment + Metadata

Rationale: Soil sample 6, a sample taken from near the bear habitat was enriched and tests for metadata were conducted in an attempt to isolate phage and gather data on the soil.

Procedure

• The lab table was cleaned with CiDecon and 70% ethanol and an alcohol lamp was lit to promote an aseptic environment.
• 4 mL of soil was vortexed for 15 minutes with 8 mL of LB broth and then centrifuged for 5 min at 3000 g.
• Percent sand, silt, and clay was tested by combining 4 mL soil with 8 mL DI water and 3 drops soil dispersion fluid and shaking for 45 seconds. That mixture was allowed to sit overnight to settle.
• Other group members performed the tests for percent water and pH. The tree was not measured because of poor weather conditions, but it will be measured and reported when the rain stops.
• 1 mL of the supernatant from the centrifuged soil was pipetted into 3 wells in a well plate and heated to 55 degrees Celsius for 6 minutes and then heated to 60 degrees Celsius for 5 minutes in order to kill off any bacteria existing in the mixture before arthro was to be added.
• 100 microliters of arthrobacter was added to two of the three wells (last one was left empty to serve as a control) and then the shaker thermostat was left to shake and incubate the bacteria overnight (27 degrees Celsius).

Observations, results, data

pH: 6.5

The soil was collected during a thunderstorm so the percent water will be much higher because of the rain. It was also very difficult to measure out when attempting the metadata tests, but this shouldn’t change percent sand, silt, clay much. It may have raised the pH because the soil itself was already so watered down from the rain.

The experimental procedure was adapted in this experiment because of a lack of materials, but new filters were recently delivered and the prior trials can be repeated as before.

Interpretations, conclusion, next steps

The arthrobacter/lysate mix will be filtered and prepared for PCR in order to amplify any present phage DNA so that it can be tested for presence of phage DNA.