Alignment of the amino acid sequence of TaRab7 and selected Rab7 proteins.

<p>Alignment of the amino acid sequence of <i>Triticum aestivum</i> Rab7 (<i>TaRab7</i>) with other Rab7 proteins from <i>Oryza sativa</i> (<i>OsRab7</i>), <i>Hordeum vulgare</i> (<i>HvRab7</i>), <i>Arabidopsis</i> (<i>RabG3b</i>), <i>Homo sapiens</i> (<i>HsRab7</i>) and <i>Saccharomyces cerevisiae</i> (<i>ScYpt7</i>). Sequences were aligned using DNAMAN. Identical residues in all organisms are shaded. Red underlines indicate sequence motifs involved in nucleotide binding and hydrolysis that are conserved in Rab GTPases.</p

Histological observations during the incompatible interaction between wheat and the stripe rust fungus when the transcription of <i>TaRab7</i> was repressed.^a

a<p>Abbreviations: hpi.: hours post inoculation.</p>b<p>Distance from the base of substomatal vesicles to hyphal tips. Values with * are significantly different at P = 0.05 according to the Tukey's test.</p>c<p>BSMV: γ and BSMV: TaRab7, leaves inoculated with BSMV: γ or BSMV: TaRab7 followed by infection with CYR23.</p

PCR primers used for <i>TaRab7</i> cloning, expression vector construction and qRT-PCR amplification.

<p>Sequences underlined are restriction sites for cloning.</p

A representative phylogenetic tree of TaRab7 and selected Rab7 proteins.

<p>A representative phylogenetic tree of <i>TaRab7</i>, Rab family member proteins in <i>Arabidopsis</i>, <i>Oryza Sativa</i> and <i>Brachypodium distachyon</i>, and Rab7 proteins from soybean (<i>GmRab7</i>) and barley (<i>HvRab7</i>). GeneBank accession numbers are provided after the gene names.</p

Assays for the transcript levels of <i>TaRab7</i> during different infection stages.

<p>Expression levels of <i>TaRab7</i> in wheat leaves inoculated with CYR23 (incompatible reaction) and CYR31 (compatible reaction) sampled at 0, 6, 12, 24, 48, 72, and 120 hpi. Relative expression was calculated by the comparative threshold (ΔΔCT) method. Mean and standard deviation were calculated with data from three independent biological replicates.</p

Relative transcript levels of <i>TaRab7</i> assayed by qRT-PCR after <i>TaRab7</i> gene silencing by the BSMV-VIGS method.

<p>RNA samples were isolated from the fourth leaves of the wheat seedlings infected with BSMV:TaRab7 at 0, 24, 48 h and 120 post-inoculation (hpi) after inoculation with CYR23. CK: wheat leaves infected with BSMV:γ were sampled immediately (0 hpi) after inoculation with CYR23. Error bars represent the variations among three independent replicates.</p

Histological observation of leaves treated with recombinant BSMV viruses and infected with avirulent race CYR23.

<p>Typical leaves were examined at 24, 48 and 120 hpi. Treatments a, b, c: BSMV:γ infected leaves inoculated with CYR23 examined at 24, 48 and 120 h, respectively. Treatment were d, e, f, BSMV: TaRab7 infected leaves inoculated with CYR23 examined at 24, 48, 120 h. SV, substomatal vesicle; IH, infection hypha; HMC, haustorial mother cell; NC, necrotic cell; SH, second hypha; HC, haustorium cell. Bars = 20 µm.</p

Assays for the transcript levels of <i>TaRab7</i> during different treatment stages.

<p>Quantitative reverse-transcription polymerase chain reaction analyses of <i>TaRab7</i> expression in wheat exposed to environmental stresses. The data were normalized using the expression level of wheat translation elongation factor 1 alpha-subunit. Mean and standard deviation were calculated with data from three independent biological replicates. High salinity, 200 mM NaCl; drought, 20% PEG6000; low temperature, 4°C.</p

Subcellular localization of TaRab7 protein.

<p>Onion epidermal cells were transformed with plasmids expressing the fusion protein and green fluorescent protein (GFP) by bombardment. All images were observed with a confocal microscope. (A). Onion epidermal cells expressing the GFP alone driven by the 35 S promoter. (B). Onion epidermal cells expression the TaRab7-GFP fusion protein.</p

Mild chlorotic mosaic symptoms were observed on the leaves at 9 dpi.

<p>Mild chlorotic mosaic symptoms were observed on the leaves inoculated with BSMV: γ or BSMV:TaRab7 at 9 dpi. Photobleaching was evident on leaves infected with BSMV: TaPDS at 15 dpi but not on mock-inoculated leaves.</p