Rationale:

The purpose of todays lab was to run a plaque assay on the newly enriched soil sample gathered previously in hopes of finding any evidence of phage presence. It was discovered that this year the Arthrobacter cultures we have been using may not have even been Arthrobacter, so for this lab we used a culture grown and kept from 2017.

Materials:

• 2.5-mL 2X TA
• 500-μL Arthobacter 
• 2.0-mL LB Broth
• 22.5-μL Calcium Chloride
• 10-μL Enriched Lysate

Procedure:

1. Established aseptic zone.
2. First began by checking soil composition results.
3. Once soil composition was recorded, began plaque assay procedure by filtering enriched lysate through 0.22 micron filter.
4. After filtration, added 2.0-mL LB Broth into a conical vial.
5. Aliquoted 22.5-μL of calcium chloride into the LB Broth.
6. Combined 10-μL of lysate with the 500-μL of Arthrobacter and left to infect.
7. Plates had to be warmed for approximately 25-30 minutes.
8. Once plates were obtained, combined lysate with the broth mixture.
9. Next added 2.5-mL of 2X TA to broth mixture and plated immediately.
10. Once solidified, plate was inserted into the incubator until next lab.
11. After incubation, grabbed a weighing plate and recorded the mass.
12. Added some of the new soil sample, recorded the mass, then left out for water to evaporate until next lab.

Results/Data:

• The soil composition had approximately 1.8-mL of sand, 1.2-mL of silt, and 0.5-mL of clay. It also had an extremely large amount of organic material floating around on the top that had to be removed. This makes a percent composition of 51.4% sand, 34.3% silt, and 14.3% clay.

• 

  Soil Composition of Sample 4 10/08/18

• The mass of the wet soil was 1.93-grams. This was gathered after subtracting the mass of the weigh plate (1.96-grams) from the total mass of the weigh plate and soil (3.89-grams).
• Plaque Assay went very smoothly and extra precautionary steps were taken to ensure an aseptic zone was maintained.

Conclusions:

• Based off the percent compositions and the soil composition chart below, the soil gathered is sandy loam. This soil is dominated by sand particles, but has enough silt and clay to allow some structure and fertility. This would make sense as the soil was sandy enough, yet was able to retain its shape and had a fair amount of water present in the soil.
• 
• Large amount of organic material present in the soil composition was due to the amount of grass that was gathered in the process of digging the soil sample as grasses were everywhere.

Next Steps:

• Analyze plaque assay results to determine presence of phage. If plaques are present, they will be picked and diluted to run plaque assays to begin purification and amplification of phage. If no plaques are present, one last soil sample will be gathered.