Forming Sustainable Communities under Increasing Environmental Constraint and Decreasing Population

Additional file 6: Fig. S5. Contact angle measurement of E. coli knockout strains (A) JM109 (control), (B) △yghW, and (C) △yibT. Three biological replicates were performed

Primers for PCR amplification of genes <i>mmsB</i>, <i>tsf, and PSEEN0851</i>.

a<p>The underlined sequences are the restriction enzymes sites.</p

Effect of organic solvents on cell growth of <i>P. putida</i> JUCT1 (open bar) and its parent strain JUCS (gray bar).

<p>The strains were initially grown in nutrient medium at 37°C till OD₆₆₀ reached 0.2, and then 60% (v/v) organic solvent was added for further incubation of 5 h.</p

Cell growth of recombinant <i>E. coli</i> JM109 strains in the presence of 4% (v/v) cyclohexane.

<p>The recombinant strains were cultured at 37°C and cyclohexane (CH) was added when OD₆₆₀ reached 0.2 (indicated by the arrow). □, pQE-80L (as the control); ▾, pQE-<i>mmsB</i>; •, pQE- <i>tsf</i>; △, pQE-<i>PSEEN0851</i>.</p

Colony formation of <i>E. coli</i> JM109 strains over-expressing <i>mmsB</i>, <i>tsf</i>, and <i>PSEEN0851</i> on LBGMg agar overlaid with decalin after 24 h incubation at 37°C. JM109 carrying empty pQE-80L was used as the control.

<p>Colony formation of <i>E. coli</i> JM109 strains over-expressing <i>mmsB</i>, <i>tsf</i>, and <i>PSEEN0851</i> on LBGMg agar overlaid with decalin after 24 h incubation at 37°C. JM109 carrying empty pQE-80L was used as the control.</p

SDS-PAGE analysis of expression of <i>mmsB</i>, <i>PSEEN0851</i>, and <i>tsf</i> in <i>E. coli</i> JM109.

<p>The transformants were grown in LB at 37°C and induced by 1 mM IPTG. Lanes: M, molecular mass marker; 1, pQE-80L (as control); 2, pQE-<i>mmsB</i>; 3, pQE-<i>PSEEN0851</i>; 4, pQE-<i>tsf</i>. Arrowheads indicate the locations of recombinant proteins.</p

2-DE images of protein extracts of <i>P. putida</i> JUCT1 grown under different solvent conditions.

<p>a: nutrient medium without solvent; b: with 60% (v/v) cyclohexane; c: magnification of c1–c3, the protein spots selected in this study are circled. Arrowheads indicate the protein spots exhibiting intensity discrepancy of over 50% in samples a and b.</p

The oxidation of cyclohexane by <i>E. coli</i> strains.

<p>Incubation condition: 37°C for 8 h with cyclohexane (1%, w/v).</p

MALDI-TOF/TOF analysis of peptides from 5 high-abundance proteins.

a<p>Number of peptides identified for individual protein in MALDI-TOF/TOF analysis.</p>b<p>Number of unique peptides identified for individual protein.</p>c<p>Total amino acid sequence of peptides identified for individual protein.</p>d<p>Percentage of amino acid sequence covered by peptides of individual protein.</p

Five high-abundance protein spots identified by MALDI-TOF/TOF.

a<p>The expression level of <i>P. putida</i> JUCT1 grown in nutrient medium without solvent.</p>b<p>The expression level of <i>P. putida</i> JUCT1 grown in the presence of 60% (v/v) cyclohexane.</p