Fig. 5
Expression of gfp using a 5′-fusion sequence from AW0309160_00174. AW0309160_00174 was by RNA-sequencing identified in this study as the most highly expressed gene under our standard growth conditions. a Picture shows cell pellets of A. wodanis containing pTM214_P174_GFP (contains 300-bp of the AW0309160_00174 gene promoter), or no plasmid (control) in microcentrifuge tubes subjected to UV light. Bright green color shows strong expression of gfp. b Secondary structure model of the first 60-nt of AW0309160_00174 mRNA. The sequence was used as a 5′-fusion to improve protein expression (gfp shown as example). c Schematic figure showing the expression cassettes of plasmids pTM214_His-GFP, pTM214_P174_GFP and pTM214_5′174_GFP. pTM214_P174_GFP contains a 300-bp region of the AW0309160_00174 promoter placed in front of gfp, and the latter contains a Ptrc promoter in front of a 60-nt/20-aa 5′-fusion from AW0309160_00174 followed by gfp. pTM214_5′174_GFP was used as backbone for cloning and expression of non-Aliivibrio enzyme test cases. c, d Fluorescence measurements of A. wodanis containing no plasmid (Control), pTM214_His-GFP or pTM214_5′174_GFP. Samples with (+) or without (−) IPTG are shown. Values are expressed as relative fluorescence units (RFU)