Home FAQs How should I calculate the transformation efficiency of NEB 5-alpha Competent E. coli (Subcloning Efficiency)?

FAQ: How should I calculate the transformation efficiency of NEB 5-alpha Competent E. coli (Subcloning Efficiency)?

Transformation efficiency is defined as the number of colony forming units (cfu) which would be produced by transforming 1 µg of plasmid into a given volume of competent cells. The term is somewhat misleading in that 1 µg of plasmid is rarely actually transformed. Instead efficiency is routinely calculated by transforming 100 pg-1 ng of highly purified supercoiled plasmid under ideal conditions. If you plan to calculate efficiency to compare cells or ligations, keep in mind the many variables which affect this metric.

Transformation efficiency (TE) equation:
TE = Colonies/µg/Dilution
Colonies = the number of colonies counted on the plate
µg = the amount of DNA transformed expressed in µg
Dilution = the total dilution of the DNA before plating

TE calculation example:
Transform 1 µl (1 µg) of 1 mg/ml pUC19 DNA into 50 µl of cells, outgrow by adding 950 µl of SOC and dilute 10 µl up to 1 ml in SOC before plating 100 µl. If you count 250 colonies on the plate, the TE is:
Colonies = 250
µg DNA = 1.0
Dilution = 10/1000 x 100/1000 = 0.001
TE = 250/1/0.001 = 2.5 x 105 cfu/µg