October
18
10/15 ~ Re-performing the plaque assay to confirm titer strength
Rationale: Re-performing the plaque assay procedure for the 10^0, 10^-1, 10^-2 and 10^-3 dilutions due to contamination of the plates. Will be using the 10^0 lysate obtained from last lab
Procedure:
- Created an aseptic zone to prevent the contamination of plates
- Obtained 10^0 lysate from fridge and obtained five plates
- Created the 10^-1 lysate by pipetting 90μL PB and 10μL 10^0 into a micropipette
- For the 10^-2, added 90μL PB and 10μL 10^-1
- For the 10^-3, added 90μL PB and 10μL 10^-2
- Added the lysates (Respectively) to 0.5mL arthrobacter
- Obtained a 50mL conical vial and pipetted 10mL Lb Broth and 112.5μL CaCl2
- Added 12.5mL 2XTA to the conical vial and immediately pipetted 4.5mL into the respective arthrobacter+lysate vials and plated
- Allowed plates to sit for 15 minutes and then incubated
- Cleaned lab bench
Observations:
- The TA plate was contaminated with spots
- The plaque assays yielded no plaques
Next Steps/Conclusion: After this plaque assay, hopefully it will yield positive results and measurable results. The next step would be to calculate the strength of the current titer, and see if it’s high enough to continue to DNA extraction