Rationale: Collected more soil, started setting up and collecting data about the soil metadata and washed the soil. Decided to get a new soil sample since plaque assay was negative. Obtained a new soil sample for soil C, washed the soil, and started on soil metadata, since one out of the three plates turned positive.  

Procedure: 

1. Recollected soil sample C and then collected new soil from a nearby tree.  
2. Filled up a 50mL tube with soil and filled up 2mL of soil in a 15mL vial. 
3. Added 10 mL of LB broth into each of soil samples and shook for 10 minutes.  
4. While waiting, started on setting up for soil metadata.  
5. Obtained a weigh boat and weighed it, then added soil and reweighed it.  
6. Repeated for Soil Sample C and placed both in the hood to be observed next class.  
7. To find percent sand, silt, clay obtained vial, and filled soil to 10 mL line, and DI water to the 30 mL line.  
8. Added soil dispersion liquid into the vial , shook if for 30 seconds, and placed it under the hood.  
9. After 10 minutes was up, massed the soil solution and placed it into the centrifuge for 10 minutes at 5,000 x g. 
10. Continued by adding a small amount of soil into a small vial, filled the rest with DI water  shook tube, let it set for 10 minutes, and placed less than a inch of PH paper inside solution to obtain pH of soil samples.  
11. After centrifuging the soil solution, the supernatant, which will be the enriched lysate, was extracted using a syringe and placed into a 50 mL vial.  
12. Added 0.5 mL of Arthro into vial and stored in a shaking device.

Observations/ Results: 

Plaque Assay:  

Plates were not contaminated, but both the control and individual plate slid, and resulted in negative plates.

Soil metadata: 

• Mass of soil D and LB broth = 18.479 grams 
• % water – weigh boat for Soil Sample D = 2.478 grams  
  • Total with soil Sample D = 6.205 grams 
• weigh boat for Soil Sample C = 2.33 grams  
  • Total with soil Sample C = 6.388 grams 
• Observed that while setting up the % sand, silt, clay soil metadata for soil Sample C, the soil started floating. 
• PH of soil C = 5.5  
• PH of soil D = 5.5  
• Total enriched lysate = 10 mL  

Next Steps: 

Next class we will collect the remaining soil metadata, specifically percent water and the percent sand, silt, and clay in the soil. Also we will be performing a plaque assay for both Soil Sample C and Soil Sample D.