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A high frequency of transformation of bacteria of about 1—5xlO cells per /tg of supercoiled plasmid DNA (1, 2) is now routinely obtained when a dense suspension of Escherichia coli (MC1061/P3; Invitrogen) cells ( lxlO 0 cells/ml) are exposed to a brief (3 -5 msec) high voltage electrical pulse. However, if the DNA sample is dissolved in a ligation mixture with a high salt concentration, the transformation frequency is decreased substantially. To circumvent this difficulty, dilution (3), dialysis (4), precipitation (5) or heat inactivation (6) of the ligation mixtures have been reported. However, none of them are entirely satisfactory. They hardly exceed the frequency of the competent cells prepared by a chemical treatment we developed recently (7), in which the DNA in a ligation mixture can be directly introduced without any treatments, giving transformation frequency of 1 3 X10 cfu per 1 y.g of pBR322 DNA. Here we report a new protocol which retains about 42% of the original high transformation frequency in a ligation mixture after a simple treatment.

A simple treatment of DNA in a ligation mixture prior to electroporation improves transformation frequency