Cell size differences in plasmid-containing and plasmid-free cells of Bacillus subtilis during batch and cyclic batch culture

Differences in cell sizes were observed in recombinant Bacillus subtilis studied in cyclic batch culture. It was observed that under this mode of operation both plasmid-containing and plasmid-free cells were present in the culture broth for an extended period of time when compared to batch culture. In cyclic batch culture, plasmid-containing cells decreased from 100% to 0% over a period of 75 h, compared with batch culture where plasmid-containing cells decreased to 0 over a period of 20 h. Plasmid copy number was found to be similar in cyclic batch culture to that found in batch culture. Copy number increased from 90 copies up to a maximum of 190 copies in cyclic batch culture after the drop in plasmid-containing cells below 40%. In batch culture, copy number ranged from 90 to 140. Cell size distributions by using image analysis and particle-size equipment indicated the existence of two populations in cyclic batch culture. Average cell size decreased with decreasing percentage of plasmid-containing cells, whereas the average plasmid-containing cell size increased with increasing plasmid copy number.