Title: Dilutions for a Flooded Plate

Date: 14 November 2018

Rationale: The webbed plates (30 uL and 10uL) webbed completely and were flooded before this lab entry. The flood lysate will be diluted and a titer calculation done in order to determine of the lysate is a high enough titer.

Procedure: Under an aseptic zone,

• The plates containing the flood lysate were filtered through a 22-micron top filter
• Once the lysate was filtered, 10 uL of the lysate (“Flood Lysate 7”) was transferred to a micro-centrifuge tube containing 90 uL phage buffer to create a 10^-1 concentration diluted lysate
  • 10 uL of the 10^-1 was then transferred to a second micro centrifuge tube containing 90 uL of phage buffer
    • This process was repeated until a 10^-4 concentration lysate was created
• Four plaque assays were then done (+ a control) to determine the titer. The following recipe was used:
  • 10 mL LB Broth
  • 12.5 mL 2x Top Agar
  • 112.5 uL CaCl2

~ 4.5 mL transferred to tube containing 0.5 mL Arthrobacter + Plate 1) 10 uL 10^-2 lysate

Plate 2) 10 uL 10^-3 lysate

Plate 3) 10 uL 10^-4 lysate

• The plates were then placed into the incubator

Results/Conclusions: Below are the results of the previous experiment. The titer of the previous flood lysate was too low to be acceptable, and the control showed a growth, but it likely resulted from a mistake in the incubator, as it was clean when checked the previous day. If the titer of the plates done in this experiment do not meet the acceptable value, more plates will be webbed and flooded.