Escherichia coli gt115

Briefly, the CD::UPRT::GFP plasmid was constructed using the vector backbone of pCpGfree-Lucia (InvivoGen). Lucia was replaced by CD::UPRT and GFP using pSELECT-zeo-FcyFur and CpG-free GFP:Sh as template. The functional plasmid of interest, CD:UPRT:GFP, was constructed using cross-lapping in vitro assembly cloning method as described [50 (link)]. The construct (Additional file 1) was transformed using heat shock method with chemically competent Escherichia coli GT115 (Invivogen) and propagated under Zeocin antibiotic selection. The plasmids were then extracted and purified with E.Z.N.A. endo-free plasmid maxi kit according to the manufacturer’s instruction (Omega Bio-tek).

Vibrio fischeri strains used in this study are listed in Table 1, and plasmids used are listed in Supplementary Table S1. V. fischeri strains were derived by conjugation. Escherichia coli GT115 (Invivogen, San Diego, CA, USA), π3813 (Le Roux et al., 2007 (link)) and S17-1λpir were used (Simon et al., 1983 (link)) for cloning and conjugation experiments (Boettcher and Ruby, 1990 (link); Visick and Skoufos, 2001 (link)). V. fischeri strains were cultured in Luria-Bertani salt (LBS) medium (Stabb et al., 2001 (link)). The following antibiotics were added to LBS medium at the indicated concentrations: chloramphenicol (Cm) 2.5 μg ml-1, erythromycin at 5 μg ml-1, and tetracycline (Tet) at 5 μg ml-1. E. coli strains were cultured in Luria-Bertani medium (LB; Davis et al., 1980 ). The following antibiotics were added to LB medium at the indicated concentrations: kanamycin (Kan) at 50 μg ml-1, Tc at 15 μg ml-1, or ampicillin (Ap) at 100 μg ml-1. For solid media, agar was added to a final concentration of 1.5%.