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From 2010.igem.org

Contents 1 Andreas 1.1 Preparation of Top10 chemically competent cells 1.2 Extraction of RBS BioBrick (BBa_B0034) 1.2.1 Colony PCR 1.2.2 Gel verification

Andreas

Preparation of Top10 chemically competent cells

Picked 8 single colonies from the 10/9 LB plate with pipette tips. Colonies were restreaked onto an LB agar plate with Amp 100 to verify that they were not contaminated, and the pipette tips were then used to inoculate cultures of 5 ml LB. Cultures grown for 40 h in 30 °C w/o rotary shaking.

Extraction of RBS BioBrick (BBa_B0034)

Colony PCR

Picked 4 colonies (RBS 1-4) from the 10/9 ON plate and ran a colony PCR.

• Positive control (PC): pSB1C3.RFP
• Standard colony PCR protocol
  • Elongation: 0:45

Gel verification

1.5 % agarose, 90 V

Expected bands

• RBS 34: 250 bp
• PC: 1385 bp