Reagants:
LB broth (Luria Bertrani medium = rich media to grow bacteria)
TSS buffer (to prepare chemically competent cells)
S.O.C. medium (helps obtain the maximal transformation efficiency)
LB agar (gel where bacteria can grow)
Chloramphenicol (CAL) at stock concentration 25mg/ml

Preparation of chemical competent cells:

1. Inoculate DH5α cells into 50mL LB and incubate at 37°C
   
2. Monitor growth every 30mins by measuring optical density at 600nm (OD600); until reach OD600 = 0.4-0.6
   
3. Harvest cells and prepare using TSS protocol (see: TSS Protocol)
   
4. Aliquot 200uL and flash freeze at -80°C

LB Agar plates preparation:

1. Prepare LB containing chloramphenicol (CAL) (at 25μg/ml)
   
• Melt LB in micorwave (defrost setting for 15mins)
• Cool LB by running cold water over
• Stock of 25mg/ml CAL → so add 400μl CAL to 400ml LB
2. Pour plates (in fume hood) and allow to solidify
   

Chemical Transformation:

1. Add 1μl of DNA to 50ul of competent cells, mix well and place on ice for at least 30mins
   
2. Heat shock cells at 42°C for 30secs, followed by 2min incubation on ice
   
3. Add 450μl of SOC medium tot he cells and incubate for 45min at 37°C (to allow (antibiotic resistance) protein expression)
   
4. Plate and spread (glass spreader sterilised over a flame and in ethanol) 50, 100, and 200μl of the cells into the agar plates made previously
   
5. Incubate at 37°C for 2h