User:DavidC/23 September 2009

Ligation
Ligation scheme: plasmid / insert:

BBa_C0012/BBa_B0014

First report: Plasmid = 0,5µL Insert = 3µL

Second report: Plasmid = 0,5µL Insert = 7µL

Third report: Plasmid = 0,5µL Insert = 1µL

Fourth report: Plasmid = 0,5µL Insert = 5µL

NEB Enzymes
For each samples add sterilized water to obtain a maximum volume equals to 8µL.

Ligation mix (NEB): 2µL of T4 ligase + 2µL of T4 ligase buffer. Add 2µL / tube. Incubation 1h at RT.

Electroporation
Electroporation cuvettes = 2mm ; inoculums of electrocompetent E.coli DH5alpha= 40µL; pulse = 2,5KVolt ; 1h of incubation.

Spread 1mL of inoculums into a petri dish with LB + ampicillin (50mg/mL) (20/0,02mL).