IGEM:Harvard/2010/General Protocols/EColi

E. coli is a typical bacteria used for molecular cloning. Long-term storage of BioBrick plasmids is in E. coli suspended in glycerol (500ul cells:500ul glycerol) and kept at -80˚C. Growing E. coli is performed at 37˚C, at which temperature they divide every 20 minutes.

• Day1: Making streaks from glycerol stocks
  • Warm agar plate at 37˚C for 10 minutes
  • Label lid of plate with what you're streaking on it, and your name, and the date
  • Locate desired glycerol stock in -80˚C freezer and, using a sterile toothpick, scrape out a tiny bit and streak on plate
  • Incubate plate overnite at 37˚C

• Day2: Growing liquid cultures
  • Label 15mL falcon tube and add 5mL of LB media + selective antibiotic
  • Using 200uL tip w/o filter, touch streak or single colony and put tip in falcon tube
  • Grow culture overnite in a shaking 37˚C incubator