Stability of plasmid pHV1431 in free and immobilized cell cultures. Effect of temperature

The segregational and structural stability of pHV1431 has been examined in Bacillus subtilis grown at 30 and 37 degrees C in continuous cultures without selection pressure. Immediately after appearance of plasmid-free cells in the reactor, a competition was observed between bacteria that favored plasmid-free cells because of the faster growth. A stronger instability was found at 30 degrees C compared to that at 37 degrees C. At 30 degrees C after 50 hours of culture, 2% of the cells carried the plasmid, whereas at 37 degrees C this percentage was reached after 130 hours. In both cases, no structural instability was observed. To improve the stability, the recombinant Bacillus subtilis (pHV1431) was immobilized in kappa-carrageenan gel beads. In comparison to free cell systems, a higher cell concentration was obtained. Moreover, the plasmid was maintained stable for longer periods; after 150 hours of culture 40% of cells in the reactor still carried the plasmid at both temperatures.