IGEM:IMPERIAL/2006/LabCalendar/2006-7-11

• IGEM:IMPERIAL/2006/LabCalendar/2006-7-11 Events
• Day in the lab
• Project Progress

Miniprep- Prepare Plasmid DNA for electrophoresis

Follow miniprep protocol to prepare plasmid DNA samples for gel electrophoresis. Protocol for bacterial preparation for digestion:

• Transfer 1.5 mL of culture into eppendorf
• Spin for 20 sec and remove supernatant
• Add 100 uL of solution 1 and vortex to resuspend
• Add 200 uL of solution 2
• Add 150 uL of solution 3
• Mix by inverting, white ppt appears
• Spin for 3 minutes
• Pour supernatant in new labeled tube, add 1 mL of ethanol and mix
• Spin for another 3 minutes and remove supernatant
• Spin to remove resitudal liquid and then resuspend in 50 uL of MiliQ (ultrapure water) (1 sec on vortex, don't resuspend pellet)
• Spin for 1 minute
• Read to digest!

Make-up of solutions required for digest preparation

Solution 1

• 90 g glucose
• 10 mL 0.5M EDTA
• 6.25 mL 2M Tris pH 8
• Make up to 500 mL

Solution 2

• 50 mL 10% SDS
• 20 mL 5M NaOH
• Make up to 500 mL

Solution 3

• 147 g Potassium acetate
• 57.5 mL glacial acetic acid
• Make up to 500 mL