Initiating a Starter Culture

1. Pick a single colony from a freshly streaked plate and inoculate a starter culture with 3 to 5 mL of media.
   
2. Use the appropriate antibiotic and incubate at 37 °C* for approximately 8 hours while shaking at 250-300 rpm.
   
3. Use the starter culture to inoculate an overnight culture. Dilute the starter culture 1:500 to 1:1000 in a large flask with the appropriate volume of media and incubate at 37 °C* for 12 to 16 hours while shaking at 250-300 rpm.

*This temperature is for standard E. coli. Some bacteria require different culture temperatures.