26 November 2018 ✷ Pick a Plaque again

Rationale: Members of group 6 adopted phage from Soil 4 from Melissa (collected 10/5/18) in order to help amplify the phage sample. A plaque was picked from the picked plaque run 11/19 because the titer was too low.

Procedure

• The table was cleaned with Cidecon and 70% ethanol and an ethanol lamp was lit.
• Plaque 1 on the plate from the prior lab session was picked and swirled into 30 µL of  phage buffer. This mixture was added to 0.5 mL of arthrobacter and allowed to sit for 15 minutes while the plates were made.
• A plate for 1 plaque assay and a control plate were made with the following concentrations and volumes:

• component	volume	final concentration
  LB Broth	4 mL	–
  2X Top Agar	5 mL	1X
  1M Calcium Chloride	45 µL	4.5 µM

• The arthro/lysate mix was combined with 5 mL of the above plate mix and the plate was poured, allowed to harden, and then inverted and incubated until Wednesday at 37 degrees celsius.

Observation, Results, Data

3 plaques were seen on the plate that was picked in today’s lab and each was picked and plated. This will help isolate a single phage type from the sample and purify it so it may be studied.

Interpretations, Conclusion, Next Steps

In the following lab periods, the presence of a webbed plate will mean that the lysate is a high titer and can be studied more closely using TEM. The current titer is very low, which may indicate a weaker phage or phage presence.