Dr. Steven Wilhelm , Samantha Coy
Melt MBBM soft agar and dispense in 3.0 mL aliquots and hold at 45ºC - 50ºC in a water bath
Concentrate Chlorella to 4.0*10
8 cells/mL at 5,000 rpm for 5 min at 4ºC and resuspend with fresh MBBM
*Concentrate so that 0.3 mL can be used per plate
Dilute virus with 50 mM Tris-HCl, pH 7.8 in 1/10 serial dilutions
*Fresh lysate contains approximately 10
6 -10
7 PFU/mL (check the titer prior to plaque assay)
*Dilute sample to have 20-200 plaque forming units (PFU) per plate
To each 3.0 mL soft agar aliquot, add 0.1 mL Chlorella virus and 0.3 mL Chlorella
*Mix by rolling between palms and pour onto MBBM plate
*Tilt the plate gently to allow the entire surface to be covered
*Allow plate to solidify
Incubate at 25ºC in continuous light for 6 days or until plaques are visible to count
*Use a sharpie to count plaques once there is a visible contrast
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