NOTEBOOK

Transformation protocol

• 1) Transform cells with 50 l of 1 µl plasmid.
• 2) Dilute plasmid NEB buffer.
• 3) Incubate the mixture of cells with the resistance plasmid for 30 minutes on ice.
• 4) After 30 minutes, bring the sample to heat shock in a water bath at 42 ° C for 60 seconds, after adding 200 l SOC medium and incubate at 37 ° C for 2 hours. (Procedures performed in 2 ml tubes).
• 5) Tagging cells transformed with plasmid resistance on solid LB containing antibiotics (chloramphenicol).
• 6) 250 L of cell solution was plated on solid medium for 16 hours in an oven at 37 ° C microbiological.
• 7) After the 16 hours incubation the counting antibiotic-resistant colonies was made (observed the efficiency of transformation).
• 8) Calculation of transformation efficiency: Efficiency of transformation (dilution factor = 15) x colony count x 105 / μgDNA.