5mL Overnights


  • 10mL culture tube Use 16mm x 160mm or 16mm x 125 mm
  • 5mL LB
  • Concentrated antibiotics
  • Single colonies on a plate (Best not to start O/N from glycerol stocks)


  • Pipet 1.5-20uL of the appropriate antibiotic to the appropriate working concentration into culture tube.
  • Add 5mL non-contaminated LB. Then add antibiotic.
  • Select single colony using a sterile instrument that has been cooled and inoculate the liquid culture.
  • Place culture tube in incubator at 37oC overnight shaking vigorously (250 rpm).


For I0500, culture overnight; then add 1mM ITPG in morning and continue culture for another 3 hours. Then miniprep.