Rationale:

Performed serial dilutions and spot titer tests to obtain the new titer of the lysate obtained from the medium titer.  

Procedure: 

1. Created an aseptic zone using Cidecon and 70% Ethanol, and used a ethanol burner.  
2. Obtained lysate from webbed and flooded plates of new titer lysate. 
3. Serial Dilutions were performed by adding 10 uL of the 10 ^ 0 lysate to 90 uL of the phage buffer into a small tube.  
4. The tube was vortexed and used to create serial dilutions from 10^-1 to 10^-8. 
5. Divided two plates and labeled –1 to –8 as well as one for control.  
6. 45 uL of CaCl2, 4 mL of LB Broth, and 1 mL of Arthrobacter were added to a 50 mL tube.  
7. 5 mL of Top Agar was added to the 50 mL tube.  
8. 5 mL of solution was pipetted into the two plates.  
9. Waited 10 minutes until the plates set.  
10. Added 5 uL of lysate into respective portions of the plate.  
11. 5 uL of phage buffer was added into the control part of the plate.  
12. The plates were left to sit for another 15 minutes and placed upright into the incubator.  

Observations/ Results: 

• Small bubbles formed in the middle of the plates while pouring.   

Next Steps/Conclusion: 

Next class we will calculate the titer of the new lysate. If a high lysate is attained, then we will try to create webbed plates and flood them to obtain lysate. Or we could continue by performing TEM using our high lysate.