PLASMID RETENTION AND GENE-EXPRESSION IN SUSPENDED AND BIOFILM CULTURES OF RECOMBINANT ESCHERICHIA-COLI DH5 ALPHA(PMJR1750)

Differences in plasmid retention and expression are studied in both suspended and biofilm cultures of Escherichia coli DH5alpha(pMJR1750). An alternative mathematical model is proposed which allows the determination of plasmid loss probability in both suspended batch and continuously fed biofilm cultures. In our experiments, the average probability of plasmid loss of E. coli DH5alpha(pMJR1750) is 0.0022 in batch culture in the absence of antibiotic selection pressure and inducer. Under the induction of 0.17 mM IPTG, the maximum growth rate of plasmid-bearing cells in suspended batch culture dropped from 0.45 h-1 to 0.35 h-1, and the beta-galactosidase concentration reached an experimental maximum of 0.32 pg/cell 4 hours after the initiation of induction. At both 0.34 and 0.51 mM IPTG, growth rates in batch cultures decreased to 0.16 h-1, about 36% of that without IPTG, and the beta-galactosidase concentration reached an experimental maximum of 0.47 pg/cell 3 hours after induction. In biofilm cultures, both plasmid-bearing and plasmid-free cells increase with time reaching a plateau after 96 hours in the absence of both the inducer and any antibiotic selection pressure. Average probability of plasmid loss for biofilm-bound E. coli DH5alpha(pMJR1750) population was 0.017 without antibiotic selection. Once the inducer IPTG was added, the concentration of plasmid-bearing cells in biofilm dropped dramatically while plasmid-free cell numbers maintained unaffected. The beta-galactosidase concentration reached a maximum in all biofilm experiments 24 hours after induction; they were 0.08, 0.1, and 0.12 pg/cell under 0.17, 0.34, and 0.51 mM IPTG, respectively.