The systems for selection of bacteria transformed with vector constructs are often based on antibiotic resistance. Due to the risk of transmission of antibiotic resistance genes to other bacteria and the generally undesirable use of antibiotics, there is the demand for safer, food grade selection systems. In this master's thesis, we attempted to set up a selection system for Lactococcus lactis, which would be based on the ability of bacterial cells to metabolize lactose. We prepared a vector construct pNZ8148_cat_lacF, which enables a simple removal of the existing chloramphenicol resistance selection marker, using the SalI restriction enzyme. Compared to the commercial plasmid pNZ8149, the lacF gene on plasmid pNZ8148_cat_lacF contains its own promoter and transcription terminator. We also examined the possibility of using a potential toxin antitoxin selection system based on ribonuclease barnase and its inhibitor barstar in L. lactis. For this purpose, we prepared a vector construct pNZDual_Ldh_Bar_Pnis_Bstar. We observed that barnase inhibits the growth of L. lactis and showed that the coexpression of barstar prevents the observed toxic activity of barnase. This has been the first attempt at heterologous expression of barnase and barstar proteins in bacterium L. lactis.
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