This assay uses a lawn of phage-susceptible E.coli embedded in a layer of agarose. This top agarose lays on a bed of standard agar. If phage is present in a clone streaked onto the top agarose lawn, the lawn will be lysed on incubation.
The phage-susceptible E.coli clone culture (at Source BioScience LifeSciences we have successfully used E.coli strains TG1, DH10B or JM101) must be freshly grown the day before making bacterial lawns. If a negative control lawn is also going to be tested, we recommend using Q358 which is phage-resistant.
For clean clone culture
Sterile 100ml conical flask LB broth without glycerol Glycerol stock of E.coli to be used as the lawn: Phage susceptible clone
Phage resistant control clone
Gilson pipette (5-50µl) and sterile tips 37°C incubator
For bottom agar PBA culture plates (CONS1001)or Genetix Q-trays (X602) depending on number of clones to be tested LB agar without antibiotic
For top agarose
LB broth without glycerol Ultrapure Electrophoresis Grade agarose (GIBCOBRL, 540-551OUB) Microwave (Matsui M183BT) Waterbath Falcon 15ml conical tubes (2097) or Falcon 50ml conical tubes (2098) Clingfilm