Construction of rbs-TetR-mRFP-LVA-tripart (L173)

Transformation

Ligation L173:

• insert: D112 (ES) rbs-TetR
• vector: D187 (EX) mRFP-LVA-tripart

Insert:vector ratio : 3:1 or 4:1

Transformation of E. coli DH5 alpha competent cells

• 100 µL of competent cells
• 5 µL of DNA (ligation products or pUC19 for positive control)
• 30 min on ice
• heat shock 45 s at 42°C
• 2 min on ice
• 0,9 mL SOC
• 1 hour at 37°C
• plating on LB + ampicilline
• overnight incubation at 37°C