Abstract
The mechanism of DNA-transfer from Escherichia coli (E. coli) Hfr donor strain AT2453 to recipient strain AB1157 during conjugation process has been investigated by liquid atomic force microscopy (AFM) in this work. Compared with other techniques such as scanning electron microscopy (SEM) and transmission electron microscopy (TEM), the most fascinating advantage of AFM as a high-resolution microscope is that it allows investigations and observations of biological species in physiological conditions. The AFM system can also be used as an ultra-sensitive force-measurement apparatus as reported earlier. By decorating anti-ssDNA (anti-single-stranded DNA) antibody onto AFM probes, we can utilize the AFM force-spectroscopy and the specific binding between ssDNAs and anti-ssDNA antibody to detect the presence of ssDNA, so as to investigate the DNA transfer mechanism during E. coli conjugation in vivo. In this study, both E. coli AT2453 and AB1157 were cultured and then immobilized on glass coverslip. The F-pilus between the E. coli AT2453 and AB1157 mating pair was imaged by an AFM probe and then dissected. Another AFM probe decorated with an anti-ssDNA antibody was applied to detect the ssDNA in the dissected area and measure the binding force between ssDNA and its antibody. As comparisons, the force-measurements were also conducted on undamaged F pilus or regions away from the dissected area. In addition, the measurements on pure ssDNAs were carried out to further confirm the value of binding force between ssDNA and anti-ssDNA antibody. According to the AFM force spectra, the transferred-ssDNA was only detected in the dissected area with a binding force of 109 ± 5 pN measured. Our results provide direct evidence that the DNA was transferred through the F-pilus channel between an E. coli mating pair during their conjugation.