Journal: The Plant Cell

Article Title: The R2R3-MYB Transcription Factors MYB14 and MYB15 Regulate Stilbene Biosynthesis in Vitis vinifera ^[W]

doi: 10.1105/tpc.113.117127

Figure Lengend Snippet: Expression of MYB14, MYB15, STSs, and Stilbene Accumulation during Grape Berry Development of V. vinifera cv Pinot Noir. (A) Columns represent MYB and STS transcript profiles in flower/skins. (B) Stilbene accumulation in flower/skins. (C) MYB and STS transcript profiles in seeds. (D) Stilbene accumulation in seeds. Data points are given as weeks from the onset of ripening (véraison; labeled with an arrow), with expression values as means of three replicate PCRs (n = 6) and error bars indicating se. Note that from 6 weeks before véraison (-6), berry skin has been separated from seeds. Flowering occurred 8 weeks before véraison. Accumulation of stilbenes was measured as methanol extracts of developing grape tissues using reverse-phase HPLC analysis. (B) and (D) show relative stilbene contents of trans-piceid in developing berries and in seeds as the mean of three different experiments, with error bars indicating se. Note that each time point during developmental series is a pool of >100 berries collected from ∼20 plants growing as described in Methods. FW, fresh weight.

Article Snippet: Analysis of Stilbenoids in Grapevine via HPLC HPLC analysis of stilbene compounds was performed on a reverse-phase HPLC device (Kontron Instruments; 322 pump system/360 autosampler/335 HPLC detector) with a Symmetry C18 column (3.5 μm, 4.6 mm × 150 mm [WAT200632]; Waters) protected by a guard column ( Czemmel et al., 2009 ).

Techniques: Expressing, Labeling

Journal: The Plant Cell

Article Title: The R2R3-MYB Transcription Factors MYB14 and MYB15 Regulate Stilbene Biosynthesis in Vitis vinifera ^[W]

doi: 10.1105/tpc.113.117127

Figure Lengend Snippet: Expression of MYB14, MYB15, STSs, and Stilbene Accumulation during Grape Berry Development of V. vinifera cv Pinot Noir. (A) Columns represent MYB and STS transcript profiles in flower/skins. (B) Stilbene accumulation in flower/skins. (C) MYB and STS transcript profiles in seeds. (D) Stilbene accumulation in seeds. Data points are given as weeks from the onset of ripening (véraison; labeled with an arrow), with expression values as means of three replicate PCRs (n = 6) and error bars indicating se. Note that from 6 weeks before véraison (-6), berry skin has been separated from seeds. Flowering occurred 8 weeks before véraison. Accumulation of stilbenes was measured as methanol extracts of developing grape tissues using reverse-phase HPLC analysis. (B) and (D) show relative stilbene contents of trans-piceid in developing berries and in seeds as the mean of three different experiments, with error bars indicating se. Note that each time point during developmental series is a pool of >100 berries collected from ∼20 plants growing as described in Methods. FW, fresh weight.

Article Snippet: HPLC analysis of stilbene compounds was performed on a reverse-phase HPLC device (Kontron Instruments; 322 pump system/360 autosampler/335 HPLC detector) with a Symmetry C18 column (3.5 μm, 4.6 mm × 150 mm [WAT200632]; Waters) protected by a guard column ( Czemmel et al., 2009 ).

Techniques: Expressing, Labeling

Journal: The Journal of Biological Chemistry

Article Title: Cloning of Glycerophosphocholine Acyltransferase (GPCAT) from Fungi and Plants

doi: 10.1074/jbc.M116.743062

Figure Lengend Snippet: Identification of ScGPCAT ( GPC1 ). Yeast extracts of strains from the yeast deletion collection were incubated with [ 14 C]GPC and 18:1-CoA. [ 14 C]G3P was added to the incubation as an internal control of enzymatic activities. A , enzyme reactions in yeast from the substrates [ 14 C]GPC and [ 14 C]G3P. The yeast-acylating enzymes are indicated. B , autoradiogram of the screening on thin layer chromatography plates showing the deletion strain lacking GPCAT activity (within the hatched square ). C , complementation of YGR149W ( GPC1 ) in the gpc1 Δ deletion strain. Empty vector as well as the deletion strain ( gpc1 Δ) are also included. Irrelevant lanes from the plate have been removed from the picture (marked by a vertical line ). D , Western blot of Gpc1p. Wild type strain bearing EV or a plasmid ( GPC1-V5 ) harboring GPC1 under the control of the galactose-inducible GAL1 promoter and containing a C-terminal V5 epitope. Cells were grown on either glucose or galactose. Equivalent amounts of protein (75 μg) were loaded onto each lane. Anti-Gpc1p-V5 primary antibody and goat anti-mouse secondary antibody were employed. The blot was visualized using an Odyssey FC imaging system. G6PD was used as the loading control.

Article Snippet: The goat anti-mouse secondary antibody (LI-COR Biosciences, catalogue no. 926-322-10) was used at a dilution of 1:10,000 in TBST containing 5% milk.

Techniques: Incubation, Thin Layer Chromatography, Activity Assay, Plasmid Preparation, Western Blot, Imaging

Journal: Frontiers in Plant Science

Article Title: Transcriptome-Wide Identification of Novel UV-B- and Light Modulated Flavonol Pathway Genes Controlled by VviMYBF1

doi: 10.3389/fpls.2017.01084

Figure Lengend Snippet: Selection of transgenic hairy root (HR) lines. Lines for microarray analysis were selected under the following criteria: flavonol accumulation as visualized by fluorescence microscopy on in situ diphenylboric acid-2- aminoethylester (DPBA) stained root cross sections (A) , flavonol content as shown by high performance liquid chromatography (HPLC) analysis (B) and gene expression analysis to verify transcript levels of the VviMYBF1 transgene and its well-characterized target VviFLS1 (C,D) . DBPA staining analysis shown in (A) was conducted with sections of the VviMYBF1 line 33 (left) and control line 77 (right). HPLC results in (B) are the mean values of three independent extractions from the same biological material. Gene expression analysis in (C,D) is shown relative to the expression of UBIQUITIN1 gene (previous TIGR database id: TC32075, VIT_16s0098g01190 ) with standard deviations derived from three qPCR runs with three technical replicates each. Stars indicate significant differences ( p < 0.05) between MYBF1 and control HRs based on t -test.

Article Snippet: High performance liquid chromatography analysis was performed on methanolic extracts of HR lines using a reverse phase HPLC (Kontron Instruments 322 pump system/360 autosampler/335 HPLC detector; Kontron) with a Symmetry C18 column (3,5 μm, 4,6 mm × 150 mm, catalog no. WAT200632, Waters) protected by a guard column ( ).

Techniques: Selection, Transgenic Assay, Microarray, Fluorescence, Microscopy, In Situ, Staining, High Performance Liquid Chromatography, Gene Expression, Control, Expressing, Derivative Assay

Journal: MicrobiologyOpen

Article Title: Biological effects of paenilamicin, a secondary metabolite antibiotic produced by the honey bee pathogenic bacterium Paenibacillus larvae

doi: 10.1002/mbo3.195

Figure Lengend Snippet: Contribution of paenilamicin to the antibacterial and antifungal activity of P. larvae culture supernatants. P. larvae ERIC II wild-type strain DSM25430 and mutant strain DSM25430 Δ pam A lacking paenilamicin production were grown in MYPGP medium at 30°C. Activity of culture supernatants against different bacteria ( Paenibacillus alvei , Bacillus subtilis , B. licheniformis, and B. megaterium ) and fungi ( Saccharomyces cerevisiae , Pichia pastoris , and Fusarium oxysporum ) was tested by agar diffusion assays. Neg. control: MYPGP medium.

Article Snippet: Paenibacillus alvei , Bacillus subtilis, Bacillus licheniformis, Bacillus megaterium , and Saccharomyces cerevisiae were obtained from the DSMZ (Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH, Braunschweig, Germany).

Techniques: Activity Assay, Mutagenesis, Bacteria, Diffusion-based Assay, Control

Journal: MicrobiologyOpen

Article Title: Biological effects of paenilamicin, a secondary metabolite antibiotic produced by the honey bee pathogenic bacterium Paenibacillus larvae

doi: 10.1002/mbo3.195

Figure Lengend Snippet: Antibacterial, antifungal, and cytotoxic activity of purified paenilamicin. (A) Isolated paenilamicin (left column) showed antibiotic activity against B. megaterium, B. licheniformis, P. alvei, P. pastoris, and F. oxyporum . HPLC solvent served as negative control. (B) Increasing concentrations of purified paenilamicin were tested in agar diffusion assays against B. megaterium (0.3–20.0 μ g Pam) and F. oxysporum (5.0–40.0 μ g Pam) as bacterial and fungal indicator strain, respectively. Water was used as a control. (C) Tn5 cells were incubated for 40 h in cell culture medium (SF900 II) or in cell culture medium supplemented with HPLC solvent as negative control, cell lysis buffer as positive control, and paenilamicin isolated from DSM25430 wt. Quantitative measure of cell vitality obtained by the MTT test is shown in the bar graph. Cells incubated with paenilamicin showed a significantly decreased vitality compared to the two negative controls. Bars represent mean values ± SD of three replicates, analyzed by Student's t -test (** P < 0.01). HPLC, high-pressure liquid chromatography.

Article Snippet: Paenibacillus alvei , Bacillus subtilis, Bacillus licheniformis, Bacillus megaterium , and Saccharomyces cerevisiae were obtained from the DSMZ (Deutsche Sammlung von Mikroorganismen und Zellkulturen GmbH, Braunschweig, Germany).

Techniques: Activity Assay, Purification, Isolation, Solvent, Negative Control, Diffusion-based Assay, Control, Incubation, Cell Culture, Lysis, Positive Control, High Performance Liquid Chromatography