Preparing a phage stock

Picking a single plaque

Add 500 µL SMG and 25 µL chloroform to a 1.5-mL tube. Cut off the end of a blue (1-mL) pipette tip to make the bore wider and use it to core a single plaque from a plate (stab the tip vertically, then tilt the tip lift the cored plaque out of the plate). Transfer it to the tube (be careful when blowing the cored plaque out, as droplets can splash out of the tube). Flicker the tube and place it in the fridge for a couple of hours (overnight), to allow the phage particles to diffuse out of the plaque. A single plaque of phage λ yields about 10⁶ phage particles.

Confluent lysis

Plate between 10⁴ and 10⁵ phage on soft agar (even 10⁶ if they make small plaques). Incubate at 37 °C for about 6 hours (or until confluent lysis occurs). Add 5 mL SMG to the plate, seal with parafilm (to prevent accidental spilling) and keep it in the fridge overnight. Next morning, collect the SMG from the plate, add 50 µL chloroform and centrifuge at maximum speed for 5 min. Transfer the supernatant to a fresh tube and add again 50 µL chlorofrom. If things work nicely, you should get a titer of more than 10¹⁰ pfu/mL.

If you do it the first time, it can be helpful to plate several concentrations as well as only bacteria. The latter helps to judge whether lysis occurs.

Phages collected this way can be kept in the fridge for extended periods of time.