Rationale:

To extract from DNA from the phage “Pippa” in order to analyze it and to image my TEM grid with “Pippa” on it.

Procedures:

1. Setup an aseptic zone.
2. Added 10mL of “NMN CEW Lysate 10^0 11.14.18” to a vial labeled “DNA Extraction”
3. Added 40µL Nuclease Mix to the vial and mixed.
4. Added 4mL Phage Precipitant Solution to the same vial.
5. Incubated on shaker at 37˙C for 30 mins.
6. Let sit for 40 minutes at room temp.
7. Centrifuged at 20rpm for 20 minutes.
8. Centrifuged at 7500rpm for 20 minutes.
9. Poured off the supernatant from the pellet and froze the pellet.
10. Imaged the phage “Pippa” via TEM.

Observations:

The vial “DNA Extraction” was erroneously centrifuged at 20rpm for 20 minutes instead of the necessary 7500 rpm for the same amount of time, the proper centrifugation was conducted immediately following the erroneous one. The imaged phage was approximately 26nm  in capsid diameter and 112nm in tail length.

Conclusions/Next Steps:

The next step will be to further prep the pelleted phage for DNA extraction which can then be analyzed for the entire genome of the phage “Pippa”.