11/19/2018

Objective:

• Extract lysate from the flooded plate
• Do a serial dilution for each lysate
• Make spot tests

Procedure:

1. On 11/16/18, three plaque assay plates were flooded with 8 ml of phage buffer each. The plates were then placed in the fridge
2. The phages buffer with phages from the plate was extracted and filtered from each plate.
3. 100 μl of the lysate from each plate was transferred to three microcentrifuge tubes.
4. 10 μl of lysate from each microcentrifuge tube was transferred to another microcentrifuge tube with 90 μl buffer.
5. 10 μl of lysate from each diluted microcentrifuge tube was transferred to another microcentrifuge tube with 90 μl of phage buffer.
6. 8 ml of LB broth is transferred to a conical vial.
7. 90 μl of CaCl2 was added to the vial as well.
8. 10 ml of 2X TA was added to the conical vial.
9. 4.5 ml of the Top Agar mixture was added to each of the 3 ,0.5 ml arthrobacter test tubes.
10. The mixture is then plated on 3 agar plates. 4.5 of the Top Agar mixture was added to another plate for a control.
11. Divide the three plates with arthrobacter into 4 sections.
12. After Agar has solidified, spot each plate with the dilutions of the respective lysates and phage buffer.
13. The plates were allowed to absorb the spots for 10 minutes and were then placed in the incubator.

Analysis And Conclusion:

the plates were flooded so that we can have more lysate. the spot tests will now be used to calculate the titer of the lysate on hand.