Additional file 3: Figure S3. of RsmW, Pseudomonas aeruginosa small non-coding RsmA-binding RNA upregulated in biofilm versus planktonic growth conditions

PA4570 is unable to complement for RsmA or CsrA in biofilm production, swarming and glycogen synthesis. Biofilm, swarming, and glycogen synthesis assays of various P. aeruginosa and E. coli mutants. (DOCX 879 kb

Acanthopanax hypoleucus Makino

原著和名: ウラジロウコギ科名: ウコギ科 = Araliaceae採集地: 山梨県 鳳凰山麓 (甲斐 鳳凰山麓)採集日: 1971/9/19採集者: 萩庭丈壽整理番号: JH033824国立科学博物館整理番号: TNS-VS-98382

Additional file 1: Figure S1. of RsmW, Pseudomonas aeruginosa small non-coding RsmA-binding RNA upregulated in biofilm versus planktonic growth conditions

RsmW RNA levels increase in P. aeruginosa when grown at 37 °C versus 28 °C. Quantitative RT-PCR. (DOCX 67 kb

Genome-wide search reveals a novel GacA-regulated small RNA in species-1

Nt) and PAO1-rpoS (mutant) with a double-stranded DNA probe prepared with primers 1698F (PAO1) and 1698R (PAO1) (additional file ). RNA preparations were obtained from PAO1 (wild-type) in exponential phase (OD≅ 0.8) and stationary phase (OD≅ 4.9); from PAO6281 () in exponential phase (OD≅ 0.7) and stationary phase (OD≅ 4.8); from PAO1-rpoS () in exponential phase (OD≅ 0.7) and stationary phase (OD≅ 5.0).. Hybridization cross-linked total RNA from CHA0 (wt), CHA89 (mutant) and CHA815 (mutant) with a double-stranded DNA probe prepared with primers 1698F (CHA0) and 1698R (CHA0) (additional file ). RNA was extracted from CHA0 (wild-type) in exponential phase (OD≅ 0.5) and stationary phase (OD≅ 6); CHA89 () in exponential phase (OD≅ 0.6) and stationary phase (OD≅ 6); CHA815 () in exponential phase (OD≅ 0.6) and stationary phase (OD≅ 5.0). Hybridization of the same RNA preparations from as in , with an RsmZ probe synthesized with primers RsmZF (CHA0) and RsmZR (CHA0) (additional file ). As a loading control, 5S rRNA was revealed in all samples with a probe synthesized with primers 5S rRNA-1 and 5S rRNA-2 (additional file ). E: exponential phase, S: stationary phase.<p><b>Copyright information:</b></p><p>Taken from "Genome-wide search reveals a novel GacA-regulated small RNA in species"</p><p>http://www.biomedcentral.com/1471-2164/9/167</p><p>BMC Genomics 2008;9():167-167.</p><p>Published online 13 Apr 2008</p><p>PMCID:PMC2375449.</p><p></p

Genome-wide search reveals a novel GacA-regulated small RNA in species-3

E determined in the wild-type PAO1 (squares) and in the mutant PAO6281 (triangles). . β-Galactosidase activities of an -transcriptional fusion (pME7234) were determined in the wild-type CHA0 (squares) and in the mutant CHA89 (triangles). β-Galactosidase activities of a chromosomal '-'translational fusion were determined in a wild-type context (CHA207) carrying the empty pME6032 vector (filled squares) or the overexpressing plasmid pME7236 (open squares), and in the triple mutant (CHA1145) carrying pME6032 (filled diamonds) or pME7236 (open diamonds). 1 mM IPTG was added when the cultures were inoculated. Each value in , and is the average from three different cultures ± standard deviation. Verification of the overexpression of RgsA sRNA by Northern blot. Total RNA was purified from cultures of CHA0 (wild type) and CHA815 (mutant) grown to stationary phase, carrying the empty pME6032 vector (-) or the overexpressing plasmid pME7236 (+).<p><b>Copyright information:</b></p><p>Taken from "Genome-wide search reveals a novel GacA-regulated small RNA in species"</p><p>http://www.biomedcentral.com/1471-2164/9/167</p><p>BMC Genomics 2008;9():167-167.</p><p>Published online 13 Apr 2008</p><p>PMCID:PMC2375449.</p><p></p

Genome-wide search reveals a novel GacA-regulated small RNA in species-0

Exponential phase (first lane in each blot) and stationary phase (second lane in each blot). Experimental conditions are described in Methods. The blots for the annotated sRNA genes encoding 4.5S RNA (in IgR 888), RNase P RNA (in IgR 2510) and PrrF1, 2 (in IgR 2667) are not shown. The approximate positions of RNA standards are shown on the left. The main transcripts are pointed out by arrows. A tRNAgene is embedded in the IgR 1466; its transcript is indicated by an asterisk.<p><b>Copyright information:</b></p><p>Taken from "Genome-wide search reveals a novel GacA-regulated small RNA in species"</p><p>http://www.biomedcentral.com/1471-2164/9/167</p><p>BMC Genomics 2008;9():167-167.</p><p>Published online 13 Apr 2008</p><p>PMCID:PMC2375449.</p><p></p

Genome-wide search reveals a novel GacA-regulated small RNA in species-4

Redicted RsmA-binding motif (ANGGA) at position 46–50 in an unpaired region is highlighted by grey circles.<p><b>Copyright information:</b></p><p>Taken from "Genome-wide search reveals a novel GacA-regulated small RNA in species"</p><p>http://www.biomedcentral.com/1471-2164/9/167</p><p>BMC Genomics 2008;9():167-167.</p><p>Published online 13 Apr 2008</p><p>PMCID:PMC2375449.</p><p></p

IFN-γ activation in the presence and absence of GM-CSF did not affect macrophage survival or <i>P</i>. <i>aeruginosa</i> growth.

<p>Macrophages were plated in X-Vivo 15, treated with IFN-γ in the presence and absence of GM-CSF for 48 h and infected with PAO1-L for 4 h. No significant differences were observed in the percentage of extracellular <b>(A)</b>, cell-associated <b>(B)</b>, and total <b>(C)</b> bacterial growth among cultures of differentially activated infected macrophages. The cell-associated fraction was found to be increased in certain donors. n = 6 to 10 for all bacterial growth data. <b>D</b>. Levels of LDH in infected macrophage supernatants were not significantly affected by macrophage activation, n = 6 to 10. Significance was calculated by one-way ANOVA with Tukey’s post test. Unt: untreated macrophages.</p

Production of IFN-γ and IL-17A in CF and control PBMCs in response to several stimuli.

<p>PBMCs were stimulated with 2% (v/v) phytohaemagglutinin (PHA), 10 ng/ml staphylococcal enterotoxin B (SEB), or two concentrations of PAO1-L lysates—PA Hi (6,250 ng/ml total protein) and PA Lo (24 ng/ml total protein)—for 6 days, re-stimulated with 15 ng/ml phorbol 12-myristate 13-acetate (PMA) overnight, and supernatants collected on day 7 for cytokine analysis. <b>A</b>. CF patient PBMCs produced less IFN-γ than healthy control PBMCs in response to PHA and SEB, but not PAO1-L lysates. <b>B</b>. CF patient PBMCs produced less IL-17A than healthy control PBMCs in response to PHA, SEB and PAO1-L lysates. Graph depicts 5–95 percentile with median; healthy controls: n = 13, CF/Chronic PA and CF/Intermittent-Free PA: n = 15 each. Significance calculated by Kruskal-Wallis test with Dunn’s post test; * = p≤0.05, ** = p≤0.01, *** = p≤0.001.</p

Production of chemokines and cytokines by human macrophages upon infection with <i>P</i>. <i>aeruginosa</i>.

<p>Supernatants from uninfected and infected macrophages (MOI = 1) were collected at 2, 4, and 6 hpi and tested for the presence of IL-8, MCP-1, and MIP-1α <b>(A)</b> TNF-α, IL-6 and IL-10 <b>(B)</b> and IL-1β and IL-18 <b>(C)</b>. IL-8 and MCP-1 were detected in supernatants from uninfected macrophages. All cytokines and chemokines tested were upregulated upon PAO1-L infection, with most peaking at 4 hpi. Data presented are mean ± SD for n = 4 for all cytokines except IL-8 and IL-18 where n = 2. Significance calculated by repeated measures one-way ANOVA with Tukey’s post test; * = p≤0.05, ** = p≤0.01, **** = p≤0.0001.</p