SAMPLE PREPARATION FOR SDS-PAGE

1. Grow cultures in a defined array on petri plates of EC agar or L-agar, or if liquid media is to be used, on L-broth or other appropriate medium.
   (In the following steps: use the ones subscripted with "s" for solid media; and "b" for broth cultures.)

2. • s. Put 400 μL water into Eppindorf tubes
   • b. Centrifuge down the bacteria at 10,000 rpm for a minute or so. Then dump off the supernate (there are times that you might want to save the supernate in order to see if your component is lost to the medium).

3. • s. Using inoculating loop, transfer part of a colony on the plate to the Eppindorf tube; swirl loop to detach bacteria.
   • b. Add 400 μL of highly purified water to the centrifuge tube, vortex to disperse the bacteria.

4. After doing all colonies for the day, continue.

5. • sb. Add 400 μL of 2X-Lysing Buffer; immediately mix.
   • b. Dump the blue contents of the centrifuge tube into an Eppindorf tube.

6. Boil all the tubes for 15 minutes (Denatures proteins)
   (or autoclave for 1 minute with slow exhaust).

7. Cool

8. Samples are now ready for electrophoresis. (Such prepared samples last for months at room temperature.)