Phage-Mediated Genetic Manipulation of the Lyme Disease Spirochete Borrelia burgdorferi

This protocol is significant because it describes another important tool for dissecting the molecular biology of the Lyme disease agent Borrelia burgdorferi. The main advantage of this technique is that by using phage transduction, it provides an alternative method for introducing DNA in the Borrelia burgdorferi without requiring the application of an electric pulse, as in electroporation. This method could be used to provide further insight into the molecular mechanisms used by Borrelia burgdorferi to persist in its enzootic cycle and ultimately cause Lyme disease in humans.

To begin, inoculate 150 microliters of the appropriate Borrelia burgdorferi clone into 15 milliliters of BSK in tightly-capped sterile conical centrifuge tubes for the transduction protocol. Then, supplement the medium with the appropriate concentration of antibiotics or a combination of antibiotics for the selection and maintenance of heterologous DNA within the Borrelia burgdorferi clone and incubate the sample at 33 degrees Celsius. After the culture has grown up, centrifuge the appropriate volume of culture at 6, 000 G for 10 minutes.