Research Question:

To find out how the presence of bacteriophages in the soil around red or white oak trees has a correlation with the health condition of oak trees.

Rationale:

A Plaque Assay helps us determine if there is a presence of bacteriophages by adding Arthrobacter directly to the lysate. We can tell the existence of bacteriophages by checking the presence of plaques on the agar plate. A PCR run can identify if there are phage DNA present in the enrichment lysate, an accurate way to detect phages through their specific conserved gene that decides the length of their tails.

PCR on Sample (10.1.18):

Materials:

1. Sample DNA from enrichment filtered lysate sample (10.1.18) and groupmate sample.
2. 2X PCR mix
3. ddH2O
4. Primers (AK, AL, AM, AN, AO, AP, AQ, AR, AS, AT, AU, AV)
5. Thermocycler
6. Microcentrifuge tubes
7. Centrifuge
8. Control DNA

Procedure:

1. Added 6.5 ul of ddH2O to the 12.5 ul PCR mix repeat for three tubes.
2. Added 4ul of primers 1, 2, and 3 to three different tubes.
3. Added 1ul of both samples, at a total of 2ul.
4. Place the mix in the thermocycler for PCR.

Observations, Results & Data:

The plaque assay showed two small clearings, the shapes are irregular, and the areas are about 0.1 mm^2, and bumps on the clearings were observed.

Interpretations & Conclusions:

The clearings resemble the clearings on the last plaque assay run, and this time the clearings were more irregular and less in numbers, the possibilities are that either this is not a plaque but rather just cracks on the agar or phages with an extremely low titer.

Next Step:

The next step would be running a DNA gel electrophoresis to determine if there is DNA present in the sample lysates.