Title: DNA Extraction Setup

Date: 28 November 2018

Rationale/Past Results: The spot titer yielded a growth of one plaque on the 10^-7 spot. This means that the official titer is 1.0 x 10^9. The lysate will then be archived and prepared for DNA extraction. Enzymes and a precipitant will be added to the mixture and it will be incubated in various conditions as well as centrifuged in order to obtain a pellet of phage DNA. Below is a picture of the spot titer result:

Procedure: 

• 2.8 mL of high titer lysate was aseptically transferred to a 15 mL conical vial for archiving.
• 10 mL (x4) was transferred to 4 different 50 mL conical vials in order to prepare the lysate for DNA extraction
• The following was added to the 10 mL of high titer lysate:
  • 40 uL combined DNase 1 + RNase A
  • 4 mL phage precipitant solution
• The mixture was then incubated at 37 degrees Celsius for 30 minutes
• The mixture was then removed from the incubator and left at room temperature for 45 minutes
• The mixture was then spun in a centrifuge for 20 minutes at 10,000 xg
• The liquid was decanted and the resulting pellets dried and placed into a micro centrifuge tube to be extracted.

Conclusions: This process produces a pellet that contains phage DNA that will be extracted. THe extracted DNA will then be able to be analyzed and characterized in the future.