Additional file 1 of Zinc glycine chelate ameliorates DSS-induced intestinal barrier dysfunction via attenuating TLR4/NF-κB pathway in meat ducks

Additional file 1: Fig. S1. Procedure for intestinal permeability of FITC-D intragastric administration in meat ducks. Table S1. Effects of Zn-Gly on growth performance of meat ducks at 14 d

Identification of the polyclonal antibody against pFTO.

<p>(A) ELISA of the antiserum against pFTO; (B) Western blot analysis of anti-FTO antibody. Lanes 1 and 2: the purified pFTO protein detected by polyclonal antibody against pFTO.</p

Effect of pFTO on the proliferation of porcine intramuscular preadipocytes.

<p>Porcine intramuscular preadipocytes were seeded in a 24-well plate at a density of 1 × 10⁴ cells/well. When the cells reached about 50% confluence, 0.5 μg of the plasmid pcDNA3.1(+)-pFTO or the empty vector pcDNA3.1(+) was transfected. (A) Cell proliferation was evaluated by EdU proliferation assay after 24 h of transfection. The Click-it reaction revealed EdU staining (red). The cell nuclei were stained with Hoechst 33342 (blue). (B) The percentage of EdU-positive cells was quantified. Results are presented as mean ± SE (n = 8). ***<i>P</i> < 0.001 as compared with the control group.</p

Subcellular localization of pFTO in porcine intramuscular preadipocytes using an indirect fluorescent immunocytochemical technique.

<p>(A) Endogenous pFTO protein in porcine intramuscular preadipocytes stained first with the anti-pFTO polyclonal antibody and then with FITC-conjugated goat anti-mouse IgG (green). (B) The nucleus marked with DAPI (blue). (C) The merged image.</p

Expression pattern of <i>pFTO</i> mRNA during porcine intramuscular preadipocytes differentiation.

<p>RNA was extracted from the differentiating porcine intramuscular preadipocytes on the days 0, 3, 5, 7 and 9. <i>pFTO</i> mRNA expression was analyzed by real-time quantitative PCR. Data were the mean and SE from three independent experiments. *<i>P</i> < 0.05, ***<i>P</i> < 0.001.</p

List of genes, primer sequences, GenBank accession numbers, and product sizes in this study.

<p>List of genes, primer sequences, GenBank accession numbers, and product sizes in this study.</p

Tissue distributions of pFTO.

<p>(A) Relative expression levels of <i>pFTO</i> mRNA in different tissues. (B) Western blot analysis of pFTO protein levels in different tissues. The amount of <i>pFTO</i> was normalized to the amount of <i>β-actin</i>. Bars presented the means ± SE (n = 3).</p

Identification of porcine intramuscular preadipocytes.

<p>Porcine intramuscular preadipocytes were characterized by Pref-1 immunofluorescent staining (100×).</p

The <i>K_m</i> value of opPPL and commercial PPL against p-NPP determined by Lineweaver-Burk method.

<p>(A) The <i>K_m</i> value of opPPL. (B) The <i>K_m</i> value of commercial PPL.</p

Effect of metal ions on opPPL and commercial PPL activity.

<p>(A),(B),(C) and (D) represent the effect of Zn²⁺, Ca²⁺, Fe³⁺ and Cu²⁺ on the activity of opPPL and commercial PPL, respectively. y means relative enzyme activity; x means concentration of the metal ions; These assays were performed as described in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0114385#s2" target="_blank">Materials and methods</a> using 10.0 mM p-NPP as substrate(n = 3). The maximum value was taken as 100%.</p