Title: Titer Dilutions

Date: 5 November 2018

Rationale: The plate from the previous experiment lysed completely and the bacteria began to regrow (picture in results/observations section). Dilutions will be done of the lysate and plaque assays will be run in order to determine the titer of the lysate.

Procedure: Under an aseptic zone,

• 10 microliters of the flood lysate was transferred into a microcentrifuge tube containing 90 microliters of phage buffer. The resulting lysate was marked as “10^-1” to represent a 10^-1 serial dilution.
• 10 microliters of the 10^-1 lysate was then transferred into another microcentrifuge tube containing 90 microliters of phage buffer. The resulting lysate was then marked as “10^-2” to represent a 10^-2 serial dilution.
• This process was repeated until a 10^-6 dilution was made and 6 different lysates were available.

Six plaque assays + a negative control were run in order to determine titer, and the following recipe was used:

• 14 mL LB Broth
• 157.5 microliters CaCl2
• 17.5 mL 2x Top Agar

~4.5 mL transferred into test tube containing 0.5 mL Arthrobacter + 10 microliters of 10^-1/-2/-3/-4/-5/-6 (depending on plate label). The plates were set for 15 minutes and placed into the incubator to grow.

Results/Observations: The following picture is a picture of the plates from the previous experiment:

Conclusions: The plates from the previous experiment likely lysed completely because they were left in the incubator for too long, so the results are inadmissible. In the next lab, the dilutions will be inspected and a titer calculated.