Journal: Nature Communications

Article Title: Temporal gene regulation enables controlled expression of gas vesicles and preserves bacterial viability

doi: 10.1038/s41467-025-67667-8

Figure Lengend Snippet: Schematic representation of the genetic constructs of the dual-inducer system ( a ) and two single‑reporter constructs ( b ) for fluorescent reporter proteins expression. c , d Fluorescence fold change of dual‑inducer vs. single‑reporter across inducer concentrations. c mCherry-only vs. mCherry-dual: NS at 0 µM IPTG; P = 0.0003, <0.0001 ( P = 0.00000058), and <0.0001 ( P = 0.000003) at 20, 200, and 400 µM. d sfGFP-only vs. sfGFP-dual: NS for all. e , f Fluorescence fold change of the dual-inducer vs. single-reporter without the corresponding inducer. e mCherry-dual vs. mCherry-only. P = 0.00461, 0.0165, <0.0001 ( P = 0.00002) at 0, 50, 500 ng/mL aTc. NS at 1000 ng/mL. f sfGFP-dual vs. sfGFP-only. NS at 0, 20, 200 µM IPTG; P = 0.00461 at 400 μM. Schematic representation of the dual-inducer system induced with varying concentrations of IPTG at fixed aTc concentration ( g ) or with varying concentrations of aTc at fixed IPTG concentration ( j ). Fold change in mCherry ( h ) and sfGFP ( k ). mCherry: 0 vs. 500 ng/mL aTc across IPTG concentrations: NS at 0, 20 µM; P = 0.0238, 0.0044 at 200, 400 µM. sfGFP: 0 vs. 200 µM IPTG across aTc concentrations: NS at 0, 50 ng/mL; P = 0.0089, 0.0012 at 500, 1000 ng/mL. Fold change in sfGFP ( i ) and mCherry ( l ). i sfGFP-dual cultures at 500 ng/mL aTc without IPTG vs. 0–400 µM IPTG: NS at 0 µM; P = 0.0009, 0.0007, 0.0010 at 20, 200, 400 µM. l mCherry-dual cultures at 200 µM IPTG without aTc vs. 0–1000 ng/mL aTc: NS at 0 ng/mL; P = 0.0478, 0.0013, 0.0145 at 50, 500, 1000 ng/mL. Fold change in c – f , h , i , k , l is plotted in arbitrary units (AU, y -axis), data representing mean ± s.d. ( n = 6 biologically independent samples). Statistics: c , d , h , k by two-tailed unpaired Welch t-test; e , f , i , l vs. grey controls by Brown–Forsythe and Welch one-way ANOVA with Dunnett T3 correction.

Article Snippet: The monomeric Cherry red fluorescent protein (mCherry) gene was obtained from Addgene (plasmid #29747), and the superfolder green fluorescent protein (sfGFP) gene was acquired from Addgene (plasmid #85492).

Techniques: Construct, Expressing, Fluorescence, Concentration Assay, Two Tailed Test

Journal: mSphere

Article Title: Analysis of Teg41 and PSMα promoter activity using a divergent fluorescent reporter plasmid

doi: 10.1128/msphere.00432-25

Figure Lengend Snippet: Dual reporter plasmid assay reveals two different activity profiles for the Teg41 and psmα promoters. ( A ) Schematic of the dual reporter plasmid (pPRB4) constructed. The psmα promoter drives expression of sGFP (green), while the Teg41 promoter controls expression of mCherry (red). A translation initiation region (TIR, yellow) was added before the start codon of mCherry to initiate translation. ( B ) S. aureus AH1263 carrying pPRB4 was grown in TSB with chloramphenicol (10 µg/mL) at 37°C, and fluorescence intensities for both sGFP (green) and mCherry (red), as well as OD 600 (black), were monitored over 16 h. The maximum fluorescence intensity for each fluorophore was set at 100%. Data are represented as the average of three replicates ± standard deviation (SD). ( C ) Relationship between fluorescence intensity and bacterial growth for both the Teg41 (right axis) and psmα promoters (left axis). Raw fluorescence data were plotted against bacterial OD 600 . ( D and E ) Plasmid pPRB4 was phage-transduced into different S. aureus backgrounds: AH1263 (blue), NCTC-8325 (red), TCH1516 (green), Newman (purple), JE2 (orange), COL (black), Mu50 (brown), and UAMS-1 (dark blue). Strains were grown under similar conditions as in panel B. Promoter activities for Teg41 ( D ) and psmα ( E ) were calculated as described in Materials and Methods. Values were normalized to 100% for AH1263 ± SD. **** P < 0.0001 by analysis of variance corrected with Dunnett’s multiple comparison test.

Article Snippet: Superfolder GFP (sGFP), mCherry (codon optimized for S. aureus ), and the promoter sequence of psmα-Teg41 (268 bp) were synthesized by Twist Bioscience.

Techniques: Plasmid Preparation, Activity Assay, Construct, Expressing, Fluorescence, Standard Deviation, Comparison

Journal: mSphere

Article Title: Analysis of Teg41 and PSMα promoter activity using a divergent fluorescent reporter plasmid

doi: 10.1128/msphere.00432-25

Figure Lengend Snippet: Teg41 promoter activity is relatively stable in the absence of global regulatory proteins. ( A and B ) Plasmid pPRB4 was phage-transduced into S. aureus AH1263 (WT), AH1263 Teg41Δ3′, or AH1263 agrA ::ery strains, and the fluorescence intensities of sGFP ( A ) and mCherry ( B ) were recorded as proxies for psmα and Teg41 promoter activities, respectively. Fluorescence intensity is expressed as a percentage relative to the WT strain, which was set at 100%. ( C ) Teg41 expression in five transcriptional regulator mutants from RNA-seq. Results are shown as log2 fold change in expression compared to the WT strain in each experiment. Dotted lines represent log2FC >1 or <−1. ( D ) Plasmid pPRB4 was phage-transduced into S. aureus JE2 WT (blue), rot ::ery (red), hpf ::ery (green), or sarA ::ery (black), and Teg41 promoter activity was monitored as previously described. Promoter activity is expressed as a percentage relative to the JE2 WT strain, which was set at 100% ± SD.

Article Snippet: Superfolder GFP (sGFP), mCherry (codon optimized for S. aureus ), and the promoter sequence of psmα-Teg41 (268 bp) were synthesized by Twist Bioscience.

Techniques: Activity Assay, Plasmid Preparation, Fluorescence, Expressing, RNA Sequencing