Rational:

Performed another round of plaque assays to further isolate the bacteriophage in the sample

Procedure:

• Created an aseptic zone to prevent bacteria contamination
• Picked a plaque from the 9/17 plaque assay (10^0), and added it to 100μL of PB
  • This became the new 10^0 dilution for which the plaque assay would be performed with
• Added 10μL of 10^0 to 0.5 arthrobacter
• Obtained a 50mL conical vial and added 14mL LB Broth, 22.5 μL CaCl2, and 17.5 2X TA
  • Made enough TA for seven plates; two plates per member of the group, and one control plate
• Immediately added 4.5 mL TA to the 10^0 + arthrobacter tube and allowed to mix, and then plated
• Let the plates sit for 15 minutes and then incubated

Observations:

• The 10^0 had many plaques, similar to the 9/12 plate
• The 10^-1 only had one, singular plaque
• The 10^-2 only had one, singular plaque
  

  Contaminated TA control for 9/17

  

  The 9/17 plaque assay (10^-2) with one plaque (Circled)

  

  The 9/17 plaque assay (10^0) with noticeable plaques

  

  The 9/19 plaque assay plate

  

  The 9/17 plaque assay (10^-1) with one plaque (Circled)

  

  9/19 control plate for TA

   

• Plates turned out as expected, with 10^0 with the most plaques, and 10^-1 and 10^-2 having less plaques
• Contamination of the 9/17 Control TA is concerning, and may entail a contaminated LB Broth

Conclusion/Next Steps:

The lab group was able to complete the procedure quickly since it was a repeat of the procedure of Monday’s lab. The next steps would be to see the results of the current plaque assay, and repeat another plaque assay to continue to isolate the bacteriophage