Journal: BMC Microbiology

Article Title: Enhanced antifungal activity of bovine lactoferrin-producing probiotic Lactobacillus casei in the murine model of vulvovaginal candidiasis

doi: 10.1186/s12866-018-1370-x

Figure Lengend Snippet: Construction and expression of the secretion plasmid pPG612.1-BLF in L.casei . a The synthetic BLF gene fragment (2.1kp) was digested with restriction enzymes BamHI and XhoI, and ligated into the sticky end of the plasmid pPG612.1 which was also digested with the same restriction enzyme, resulting in the plasmid pPG612.1-BLF (5.6kp). b The plasmid pPG612.1-BLF was electroporated into L.casei using a BioRad GenePulser with single electric pulse (voltage, 1.5 kV; capacitance, 25 μF; and resistance, 400 Ω.). PCR amplification of the BamHI site and XhoI site of the plasmid pPG612.1-BLF which was extracted from the L.casei /pPG612.1-BLF strain resulted in 500 bp and 800 bp products, respectively. Lane 1, PCR product of XhoI site; Lane 2, PCR product of BamHI site. M, DNA maker. c BLF was detected in the supernatant and pellet of L.casei /pPG612.1-BLF culture by Western blotting, indicating the expression and secretion of BLF by L.casei /pPG612.1-BLF. Lane 1, supernatant of L.casei /pPG612.1-BLF culture; Lane 2, pellet of L.casei /pPG612.1-BLF culture; Lane 3, supernatant of L.casei /pPG612.1 culture; Lane 4, pellet of L.casei /pPG612.1 culture

Article Snippet: The BLF fragment was retrieved from the pUC57-BLF using BamHI and XhoI enzymes (New England Biolabs Inc., Ipswich, MA, USA) and ligated into the corresponding sites of pPG612.1, generating the plasmid pPG612.1-BLF.

Techniques: Expressing, Plasmid Preparation, Polymerase Chain Reaction, Amplification, Western Blot