Rationale: The last plaque assays have all failed since it failed the third round of purification, and group four’s control was once again contaminated. Preformed a plaque assay to pick from the original plate that the first plaque was picked, and passed. Picked soil from 2nd street

Procedure:

Before starting we had to create an aseptic zone to ensure that all bacteria were killed, and the working space would not contaminate our experiments.

1. Cleaned off the workspace with CiDecon and applied the table with 70% ethanol solution.
2. Wiped off the table after CiDecon was applied, same with the 70% ethanol solution, only, we let the ethanol solution evaporate.
3. We then got an ethanol burner, and our aseptic zone was created.

Soil Metadata

• Soil pH
  • added about 3mL of Soil B into a 15mL vial.
  • filled the rest of the vial with DI water.
  • shook the vial for about 10 seconds and then the vial sat for 2 minutes.
  • placed pH paper.
  • recorded the results.
  • pH is 6.5
  • 
• Percent(%) Water
  • got a petri dish and labeled it ML 10/3/18.
  • recorded the weight of the dish itself.
    • 2.39g
  • added soil on the dish.
  • recorded the weight of the soil.
    • 13.82g
  • 
  • Placed this in the hooded cabinet.
• Sand Silt Clay
  • added 4mL of Soil B into a 50mL vial tube.
  • added 12mL of DI water.
  • added three drops of the dispersion liquid.
  • shook for 30 seconds.
  • placed in the hooded cabinet
  • 

Plaque Picking

• retrieved the successful plaque assay that was performed on 9/12/18.
• added 100 microliters of PB into a centrifuge cap.
• picked a plaque from the plate.
• Filtered the plaque through the 100 microliters of PB.
• Grabbed a 15mL vial and added 2mL LB broth.
• added 22.5 microliters of calcium chloride into the 15mL vial.
• added 10 microliters of lysate into 500 microliters of Arthrobacter.
• added the lysate/Arthrobacter into the 15mL vial.
• added 2.5mL of 2XTA.
• mixed the solution, and quickly poured the solution onto a plate.

Observations:

Plaque assays from Monday’s experiment fail, and the control was negative. Picked soil from a new location, washed the solution, and the metadata for the Soil sample B.

Next steps/conclusions:

Plaque assays and spot test with new lysate. Record the metadata results from the %water and %sand, clay, and silt. Check the plates from the last plaque assay to see if any results. If there are plaques, then continue to round two of passage.