Sulbactam-mediated inhibition of TEM-1 β-lactamase increases specific yields of ampicillin-resistant plasmids

Ampicillin resistance is the most widely used selection marker in molecular biology. Most ampicillin-resistant plasmids originate from highly active TEM-1 β-lactamase cassette which causes very fast antibiotic decay and negatively affects long-term plasmid retention. The present work proposes a method of controlling the β-lactamase activity by the addition of an irreversible β-lactamase inhibitor. The positive effect of a β-lactamase inhibitor, specifically sulbactam, on plasmid retention is confirmed by much higher specific plasmid yields: 4-8 times higher titres in shake-flask and almost 3 times higher in bioreactor scales were observed. Among all commercially available β-lactamase inhibitors sulbactam provides the best combination of affordability and activity. The sulbactam addition is simple, fast and economic solution allowing to screen thousands of legacy pBR322 or pUC-derived plasmids harbouring TEM-1 β-lactamase cassette without engineering their sequence.