b #162
Description
flowchart TB
%% Styles professionnels
classDef start fill:#e3f2fd,stroke:#1976d2,stroke-width:3px
classDef stress fill:#fff3e0,stroke:#f57c00,stroke-width:2px
classDef assay fill:#f3e5f5,stroke:#7b1fa2,stroke-width:2px
classDef analysis fill:#e8f5e8,stroke:#388e3c,stroke-width:3px
A["<b>🦠 E. coli K-12 MG1655</b><br/>Stock -80°C<br/>18% glycerol<br/><i>Day -1</i>"]:::start
A --> B["LB 3mL<br/>7h 37°C 120rpm<br/>OD₆₀₀ = 0.3<br/><i>Day 0 matin</i>"]
B --> C["DM + 80mg/L glucose<br/>10mL overnight<br/>Acclimatation<br/><i>Day 0 soir</i>"]
%% BRANCH PRE-STRESS
C --> D1["<b>🌊 STRESS EAU USÉE</b><br/>DM + 5% EU filtrée<br/>2h 37°C 120rpm<br/><i>Day 1 matin</i>"]:::stress
C --> D2["<b>CTRL Stress</b><br/>DM seul<br/>2h 37°C 120rpm"]:::stress
D1 --> E["<b>🚿 LAVAGE CRITIQUE</b><br/>3× centrifugation<br/>4000g 5min saline<br/>Nt_inoculum vérifié"]
D2 --> E
%% 2x2 FACTORIEL
E --> F1["<b>A : CTRL total</b><br/>DM<br/>20 cultures 1mL"]:::assay
E --> F2["<b>B : SIM pur</b><br/>DM + Stress EU<br/>20 cultures 1mL"]:::assay
E --> F3["<b>C : DAMP + Chimie</b><br/>DM + 5% EU<br/>20 cultures 1mL"]:::assay
E --> F4["<b>D : Stress + Matrice</b><br/>DM + 5% EU + Stress<br/>20 cultures 1mL"]:::assay
F1 --> G["<b>🔬 FLUCTUATION ASSAY</b><br/>96-well deep plate<br/>24h 37°C 250rpm<br/>V₀=1mL → V₂₄ₕ=0.9mL<br/><i>Day 1 soir → Day 2</i>"]:::assay
F2 --> G
F3 --> G
F4 --> G
G --> H1["<b>15 puits/condition</b><br/>Dilutions 10⁻⁵<br/>TA agar<br/>CFU → <b>Nt</b>"]:::analysis
G --> H2["<b>81 puits/condition</b><br/>TA + Rifampicine<br/>100µg/mL<br/>Surface 100% sèche<br/><b>Distribution rᵢ</b>"]:::analysis
H1 --> I["<b>Nt = CFU × 10⁵ × V₂₄ₕ/40µL</b><br/>Moyenne ×3 par condition"]
H2 --> J["<b>Distribution mutants</b><br/>r = [0,2,0,15,38,1,…]<br/>16-20 valeurs/condition"]
I --> K["<b>Shinyflan R-package</b><br/>flan::fluctuation.test()<br/>Maximum Vraisemblance<br/>Luria-Delbrück<br/><b>m = f(rᵢ)</b>"]:::analysis
J --> K
K --> L["<b>μ = m/Nt</b><br/>par locus/génération<br/>IC 95%<br/>4 valeurs μ_A,B,C,D"]:::analysis
L --> M["<b>ANOVA 2×2 factorielle</b><br/>μ ~ Stress × Matrice<br/>TukeyHSD post-hoc<br/>p < 0.05"]:::analysis
%% Légende
M --> N["<b>HYPOTHÈSES :</b><br/>B>A : SIM induit<br/>C>A : Effet matrice<br/>D>B+C : Interaction<br/><i>5 jours total</i>"]
%% Liens publication
linkStyle 0,1,2,3,4,5,6,7,8,9,10 stroke:#1976d2,stroke-width:2px