19 September 2018 ✷ Third Enrichment + Metadata

Soil will be washed and enriched in order to help isolate a phage (if there is one) and metadata will help compare positive/negative results with other groups’ data in order to look for trends.

Procedure

• The workspace was cleaned with CiDecon and 70% ethanol and an alcohol burner was lit to promote an aseptic environment.
• 2 mL of soil was collected in a vial and roughly 10 mL of LB Broth was added and the vial was vortexed for 15 minutes to mix. The sample was then centrifuged for 10 minutes at 10,000g.
• In the meantime, the mass from the drying % Water test was massed and percent water was calculated to be 6.4% (see Day 7).
• Another vial was filled with 4 mL of soil and roughly 8 mL of DI water. 3 drops of soil dispersion fluid was added, then the sample was shaken for 30 seconds, decanted into a falcon tube, and stored until Monday. This is the test for percent sand, silt, and clay.
• pH was determined by placing a small amount of dirt in a pH tube and adding DI water, shaking to mix, and inserting a pH strip, which was compared to the test kit’s indicator chart.
• After the enrichment sample was done spinning, it was filtered with a 20 micron syringe filter and 9 mL was set aside to be enriched and 1 mL was set aside to be the direct sample. The direct lysate was refrigerated.
• 0.5 mL arthrobacter was added to the “enriched” sample and left in a shaking incubator until Monday.

Observatios/Results/Data

pH= 6.0

This sample is more acidic than the previous sample.

Interpretations, Conclusion, Next Steps

Next time, a spot test will be run in order to determine if phage is present or not.

Overall, the repetition of the same procedure three times because of two negative results has led to more confidence in the tests and hopefully the third attempt yields a positive result.

Both Group 5 and Group 6 had all negative results from the initial test location. This attempt compared the same types of trees in a different location, which limits the variables at play and only changes the research question from “near North Village” to “near the Science Building.”