10/17/18

Objectives:

• Make a plaque assay for the enriched lysate extracted from soil sample C

Pre-Lab Observations:

the plaque assays prepared on 10/15/18 seemed to have not properly solidified and had some air bubbles. So, although there seemed to be plaques on the plate, another plaque assay will be made to verify the presence of phage in the lysate.

Procedure:

1. The aseptic zone was set up.
2. Some of the enriched lysate prepared on 10/10/18 was filtered into a microcentrifuge tube.
3. 0.5 ml arthrobacter was retrieved and enriched by filtered lysate.
4. The vial was then allowed to rest on the test tube rack for 15 minutes
5. While the vial was resting, one Top Agar mixture was made for the group.
6.  8 ml of LB broth was transferred to a conical vial.
7. 90 microliters of the CaCl2  was transferred to the conical vial with the LB broth.
8. The vial was then set on the rack.
9. 10 ml of the 2X TA was added to the LB broth and Cacl2 after the sample was allowed to enrich the lysate for 15 minutes.
10. 4.5 ml of the top agar mixture was transferred to the test tube with the enriched lysate.
11. The contents of the test tube were then poured onto the agar plate.
12.  Part of the top agar mixture was poured into the top agar control plate for the group.
13. To let the top agar solidify, the plates were allowed to rest for 20 minutes.
14. The plates were placed upside down in the incubator, where they will remain for 48 hours.

Analysis and Conclusion 

Although there seemed to be plaques on the plate form 10/17/18, it seemed wise to redo the plaque assay to verify and avoid waste of time that may result from picking plaques. There was no contamination and all procedures were properly performed in the aseptic zone.