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From 2013.igem.org

Contents 1 Making samples for DNA sequencing 2 Making chloroamphenicol/IPTG plates 3 Streaked out samples of the limonene/pSB1C3 ligated plasmid 4 Made polystyrene strips 5 Overnight culture preparation

Making samples for DNA sequencing

Making chloroamphenicol/IPTG plates

• 400ml of agar
  • Boil for 3min x2
  • Boil for 2min x2
• Cool to 40C
• Add 800ul of chloroamphenicol at 25ug/ml
• Add 400ul IPTG at 0.15mM
• Makes 20 petri dishes

Streaked out samples of the limonene/pSB1C3 ligated plasmid

• Streaked samples 5.1, 10.1, 10.2 and 10.3 onto the chloroamphenicol/IPTG plates
• IPTG was used to induce the lac operon, and therefore induce limonene synthesis
• Plates were labelled using special symbols by only one team member to ensure double-blinded test

Made polystyrene strips

--> include video here

Overnight culture preparation

• To make glycerol broths, so sample can be frozen and preserved
• Use four universal tubes, add:
  • 4ml Luria Broth
  • 8ul chloroamphenicol
  • Add bacteria from the original plates (sample 5.1, 10.1, 10.2 and 10.3 into a single tube each respectively)

*Each sample was also re-streaked on a chloroamphenicol plate to maintain current stock.