Rapid protocol for preparation of electrocompetent Escherichia coli and Vibrio cholerae.

Electroporation has become a widely used method for rapidly and efficiently introducing foreign DNA into a wide range of cells. Electrotransformation has become the method of choice for introducing DNA into prokaryotes that are not naturally competent. Electroporation is a rapid, efficient, and streamlined transformation method that, in addition to purified DNA and competent bacteria, requires commercially available gene pulse controller and cuvettes. In contrast to the pulsing step, preparation of electrocompetent cells is time consuming and labor intensive involving repeated rounds of centrifugation and washes in decreasing volumes of sterile, cold water, or non-ionic buffers of large volumes of cultures grown to mid-logarithmic phase of growth. Time and effort can be saved by purchasing electrocompetent cells from commercial sources, but the selection is limited to commonly employed E. coli laboratory strains. We are hereby disseminating a rapid and efficient method for preparing electrocompetent E. coli, which has been in use by bacteriology laboratories for some time, can be adapted to V. cholerae and other prokaryotes. While we cannot ascertain whom to credit for developing the original technique, we are hereby making it available to the scientific community.

Bacterial genomics and pathogen evolution.

The availability of hundreds of bacterial genome sequences has altered the study of bacterial pathogenesis, affecting both design of experiments and analysis of results. Comparative genomics and genomic tools have been used to identify virulence factors and genes involved in environmental persistence of pathogens. However, a major stumbling block in the genomics revolution has been the large number of genes with unknown function that have been identified in every organism sequenced to date.