Rationale: In this lab, the objective was to isolate possible bacteriophages from previously collected soil samples in labs before in a lysate.

Procedure:

1. The first step of the experiment was to create a sterile environment using the aseptic technique. It is done so there is absolutely no contamination of the soil sample by preventing any additional microorganisms from entering the vials once opened.
2. First, the lab bench was cleaned with cidecon and wiped dry. Following that, the bench was wiped with 70% ethanol and left to evaporate. Finally, to create an aseptic zone, a burner was lit to create a convection current of air circulating to prevent any microorganisms in the air to fall on the bench and contaminate our work space.
3. Once the area was sterilized, LB broth was added to the soil sample up to the 35 mL mark in the 50 mL tube. This was done in the aseptic zone.
4. After the LB broth had been added, the sample was shaken for approximately 15 minutes, it was also put on the vortex machine whenever I got tired of shaking. The sample does not need to be shaken vigorously, it is more important to keep the shaking continuous during the entire duration of the 15 minutes.
5. After the 15 minutes were completed, I weighed my soil sample in its test tube so that I may find a partner with a similar weight for centrifugation of the soil samples.
6. Once I had found a partner, our soil samples were inserted into the centrifuge, and spun at 3,000 revolutions per minute to separate the solid pellet from the hopefully bacteriophage full supernatant liquid at the top of the vial.
7. The vials were then removed from the centrifuge and brought back to the classroom where the supernatant was run through a .22 micrometer filter to remove any impurities still left in the supernatant, leaving  15 milliliters of a yellow liquid behind called a lysate.
8. The lysate was then split by two methods of isolation. The direct isolation method took 5 milliliters of the lysate and put it into a 15 milliliter vial (aseptic method again) and stored in the fridge. The enriched isolation took the leftover 10 milliliters of lysate and added 0.5 milliliters of our host arthrobacter to the vial, then stored it as well.

Observations:

• The supernatant was a much lighter tone than the dark soil it came from.
• The lysate was almost yellow, completely different from the black soil that we started with. This may have to do with the addition of the LB broth at the beginning of the experiment.

Results:

• The experiment yielded two vials of lysate in the end, one with our bacterial host added into it, the other without it. The soil sample we began with had been purified and filtered to hopefully isolate a bacteriophage.

Conclusions/Next Steps

• The next steps are to examine the lysate to see if we successfully isolated a bacteriophage within our soil sample. If there were any bacteriophages present, the arthrobacter introduced in the enriched isolation should have multiplied by using the bacterial host introduced into the vial. The next steps will be to analyze the enriched and direct isolation lysates to look for the presence of bacteriophages.