Day 8 thoughts. Phagefinders.
Planned: Inspection of plated soil samples; do we have new viruses? Describe any new viruses in terms of their plaque morphologies. Pick the new bacteriophages for further propagation.
1) Examine the plaques that our new phages make. Photograph these.
2) Prep of a M. smeg culture from a single colony.
3) Pick plaques and enrich for pure virus via making high titer from a plaque.
Did: We have new phages today! We picked single viral plaques by using a pipette tip, and extracting the agar with the plaque beneath it, about 1/2 the depth of the plate. This was added into 222 uL MP buffer, in a microcentrifuge tube, and placed on a shaker at RT for 2 hours. 111 uL of the sample was then added into 1 mL buffer, and diluted, so that 10-0 10-1 and 10-2 could be plated as described. These plates were photographed to record the plaque morphologies.
Bill and AECOM media crew came in today and talked with the students about the experience, necessitating the students to think on their toes with respect to what they know about phages and Mycobacteria.
During the 2 h incubation of the new plaques with MP buffer, we photographed our plaque morphologies, and also diluted our D29 high titer (50 uL high titer prep, calculated to be 4 x 108 this past week.) at -3, -4, -5 and -6. Each student diluted this phage as a pair, and plated after incubation with either older smegmatis that had set in the room since Day 2, or newer Smegmatis that was brought into the room just today. (See photos).
Began 4 other soil samples incubating with MP buffer, on a shaker at 37°C.
Today's materials:
Pointers: