Electroporation of E. coli
Claire A. Woodall
Electroporation, originally developed as a method to introduce DNA into eukary-otic cells (7), has subsequently been extensively used for bacterial transformation (2,3). This procedure is an effective method for the transfer of DNA to a wide range of Gram-negative bacteria, such as Escherichia coli, and reports indicate that 109 electro-transformants per microgram of DNA can be achieved in this species (4,5). Electro-poration is probably the most efficient and reliable method for the transformation of E. coli strains using plasmid DNA.
When bacteria are subject to an electrical pulse, it is thought that the bacterial membrane is polarized, thus forming reversible transient pores through which DNA travels into the cell (for a detailed explanation of the mechanism of electroporation, see ref. 6). Pore formation starts as a membrane dimple, which then forms a transient hydrophobic pore; some of these become more stable hydrophilic pores (7,8). The exact changes in membrane structure that occur during electroporation are not known. In most cases, electroporation causes the membrane to rupture resulting in between 50% and 70% cell death. Therefore, to obtain an optimal number of transformants, it is often necessary to determine the number of cells killed, which is dependent on the cell type, field strength (explained below), and length of electric pulse.
To achieve a high number of transformants, it is important to consider variables such as electrical field strength, pulse length, and buffer choice. It may be worth determining what type (or shape) of pulse the electroporation apparatus can produce, although there is little difference between the transformation efficiencies obtained with different electroporation pulses (9). The most common type is the exponential waveform pulse that is the decay pattern from the discharged capacitor. Most manufacturers supply detailed information regarding the optimal conditions (including the pulse length, buffer, and voltage) for the electroporation of a wide variety of cells. The manufacturers' information booklets also contain up-to-date literature on methods for the electroporation of cells, and it is advisable to refer to these.
Electroporators usually have three variable parameters: capacitance