Abstract:

The electroporation conditions of L. paracasei HD1.7 were optimized, which provided the basis for construction of the knockout mutant strains of prcR. [Methods] The suicide, pUC18-prcR-tet, was imported to the L. paracasei HD1.7 by electrotransformation and the optimized parameters affecting high-efficent electrotransforming were studied preliminarily. After L. paracasei HD1.7 had been cultivated 8h, the recipient cells were treated by 12.5 μg/mL (final concentration) of benzylpenicillin and were to be continued 1h. They were washed by electrotransformation buffer of AEB1(0.3 mol/L of sucrose), as well as the 1.8-2.0 kv of field strength and one time of pulse, then added them in MRS culture media quickly and recovered 5 h. The high-efficient electrotransforming had been got, the suicide plasmid (pUC18-prcR-tet) was imported to the L. paracasei HD1.7 successfully, and part of the homologous recombination occurred. The optimized electroporation conditions of L. paracasei HD1.7 were obtained, which lay the foundation for studying the quorum sensing regulatory factor (prcR) of L. paracasei HD1.7.