Discovery of Gibbonz

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Phage Isolation Notes

September 4, 2017

Collecting Environmental Samples

  • I stopped at three locations on the highway between Stephenville and Granbury and collected soil samples
  • Followed Protocol 5.1: Collecting Environmental Samples
Sample 1 2 3
Date of Collection 9/4/2017 9/4/2017 9/4/2017
Sample Type soil soil soil
Location Description Fence line Pond over flow Road drainage
General Location Paluxy River Stock Tank Drainage
Specific Location 92665 80088 97471 83192 03093 84599
Sample Depth 1.5 inches 1 inch 0.5 inches
Ambient Temperature 88°F 88°F 89°F

September 6, 2017

Isolation of PHAGES from samples

  • Followed Protocol 5.2: Direct Isolation, then completed Protocol 5.3: Plaque Assay
    • I left the plate in the incubator for 48 hours. No phages were present.
  • Followed Protocol 5.5: Enriched Isolation – Day 1
    • I let the enriched culture incubate for 5 days

September 11, 2017

Continued Isolation of PHAGES

  • Continued Protocol 5.5: Enriched Isolation – Day 2
  • Followed Protocol 5.6: Spot Test
    • I let the filtrate sit on the top agar for 15 min
    • I inverted the plate for a second prior to incubation (caused filtrate to move a little)
    • Incubated for 48 hrs

September 13, 2017

Checked Spot Test #1 and Started Spot Test #2

  • No phage present for spot test number one of Enriched sample (See images)
  • Followed Protocol 5.6: Spot Test
    • Tried drying condensation on top of agar plate with Kimwipe
    • Borrowed Enriched sample from Travis #3 in quadrant 2
    • Borrowed Enriched sample from Mario #2 in quadrant 6
    • Let filtrate sit for 20 min before incubating for 48 hrs

 

 

 

 

September 15, 2017

Checked Spot Test #2

  • Spot test #2 was so contaminated that Dr. Edwards removed sample from incubator

September 18, 2017

Adopting a PHAGE

  • Adopted a plaque from Morgan’s Direct Sample (See images “JM”) using Protocol 5.4: Picking a Plaque
    Sample 3
    Date of Collection 9/4/2017
    Sample Type soil
    Location Description compost bed
    General Location Subdivison
    Specific Location 29° 34′ 36.480″ N,  98° 15′ 54.360″ W
    Sample Depth 1.5 inches
    Ambient Temperature 82°F

  • Then followed Protocol 6.2: Serial Dilutions
    • I did dilutions 10^-1 through 10^-6
    • On 10^-1 I used 90 micro-liters instead of 10 micro-liters
  • Then followed Protocol 5.3: Plaque Assay for each dilution.
    • Incubated for 48 hrs

 

September 20, 2017

Results of First Dilution and Start Second Dilution

  • Results (See images)
    • 10^-1: Lots of plaques, uncountable
    • 10^-2: Approximately 35 plaques
    • 10^-3: 5 plaques present
    • 10^-4—-10^-6: no plaques present 
  • Chose isolated plaque from 10^-3 plate using Protocol 5.4: Picking a Plaque (See images)
  • Then followed Protocol 6.2: Serial Dilutions again
    • I used 10 micro-liters on all dilutions this time
    • I did dilutions 10^-1 through 10^-6
  • Then followed Protocol 5.3: Plaque Assay for each dilution
    • Incubated for 48 hrs

September 22, 2017

Results of Second Dilution

  • Results (See images)
    • 10^-1: Lots of plaques, uncountable 
    • 10^-2: Over 100 plaques
    • 10^3: 3 plaques present
    • 10^-4—-10^-6: no plaques present