Rationale

A spot test will be conducted using the possible plaques obtained from the plaque assay on 9/17/18. More soil and soil metadata will be collected as well.

Procedure

1. Use the aseptic technique to clean the area.
2. 10o µL of phage buffer was pipetted into 3 micro test tubes. The tip of a pipet was then placed into each possible phage from the previous plaque assay and the placed back into each micro test tube. Set aside the tubes.
3. Pipet 2 mL of LB broth into a vial along with 0.5 mL of Arthrobacter, 22.5 µL of CaCl2, and 2 mL of 2X TA. The vial was poured onto a plate and the plate sat for 10 minutes.
4. After 10 minutes had passed, 5 µL of each micro test tube was micro pipetted onto each quadrant of the petri dish, along with 5 µL of phage buffer into a control quadrant. The plate then sat for another 10 minutes.
5. New soil was collected by lab partners during this time.
6. Soil washing then began by placing 2 mL of soil and 9 mL of LB broth into a vial. The vial was shaken for 10 minutes to mix.
7. The vial was then massed and centrifuged for 10 minutes.
8. While the soil was centrifuging, % water calculations began by massing a weigh boat, adding soil, and then massing the weigh boat again. The weigh boat was then placed in the fume hood for 48 hours.
9. The percent sand/silt/clay of the soil was also measured by adding 10 mL of soil to a falcon tube. The tube was filled to the 30 mL mark with DI water and 3 drops of soil dispersion liquid was added. The tube was covered and shaken for 30 seconds. After, it was placed in the fume hood for 48 hours.
10. To measure the pH, 1 mL of soil was placed into a vial and then filled with DI water. The vial was shaken for 30 seconds and then sat for 10 minutes. After 10 minutes passed, pH paper was put in the vial and the pH was measured.
11. After the tube containing soil was centrifuged, the supernatant was filtered using a syringe and syringe filter to produce 6.5 mL of enriched isolation.
12. 0.5 mL of Arthrobacter was added and the tube was incubated.

Observations

The mass of the tube before being centrifuged was 17.18 g. The mass of the weigh boat was 2.32 g and the mass of the soil and weigh boat was 6.20 g. The pH of the soil was 6. This soil contained more debris than the previous soil collected.

Conclusions

The results of the spot test will be observed on Monday. From there, we will see if the previous soil sample can be used or if the new sample will be need to be tested.