5 mL Overnight

What you will need:

• 10 mL culture tube Use 16mm x 160mm or 16mm x 125 mm
• 5 mL LB
• 5uL 1000X antibiotics
• Single colonies on a plate (Best not to start O/N from glycerol stocks)

Procedure:

• Add 5 mL non-contaminated LB. Do this first, then add antibiotic.
• Pipet 5uL of each antibiotic into culture tube.
• Select single colony using sterile toothpick or a flamed loop that has been cooled
• Place toothpick or loop in culture tube, stir.
• Remove toothpick or loop and place culture tube in incubator at 37 C overnight shaking vigorously (250 rpm).

Notes

• For I0500, culture overnight; then add 1 mM IPTG in morning and continue culture for another 3 hrs. Then miniprep.
• Ampicillin: use 10 ul, all others use 5 ul.