Rationale

Today we will conduct spot titers to calculate the titer of an unknown lysate from Lucy P.

Procedure

• Established an aseptic zone.
• 90 µL of phage buffer was aliquotted into 8 microcentrifuge tubes. 10 µL of flooded lysate was pipetted into tube #1 containing 90 µL of phage buffer. 10 µL of the contents in tube #1 was pipetted into tube #2. 10 µL of the contents in tube #2 was pipetted into tube #3. The process repeated down the line for all 8 tubes.
• 4 mL of LB broth, 5 mL of 2X TA, 1.0 mL of Arthobacter, and 45 µL of CaCl2 was combined into a vial. The 10 mL of solution was divided evenly between two plates and poured. The plates were set aside for 10 minutes to solidify.
• 5 µL of the contents in each tube was spot tested onto their designated quadrants on each plate. Phage buffer was spot tested on the control quadrant. The plate was set aside for 10 minutes and then placed into the incubator.

Observations

When pouring the plate, plate #2 possessed large amounts of air bubbles that could possibly be mistaken for plaques when analyzing.

Conclusions/Next Steps

Results will be assessed on 11/16/18 to determine the titer of the lysate.