Journal: BMC Plant Biology

Article Title: Elucidating the callus-to-shoot-forming mechanism in Capsicum annuum ‘Dempsey’ through comparative transcriptome analyses

doi: 10.1186/s12870-024-05033-4

Figure Lengend Snippet: Analysis of DEGs from callus and shoot tissue in Capsicum annuum ‘Dempsey’. ( A ) Example images with the scale bar of samples from leaf (WT), callus, and shoot tissues used for RNA-seq analysis: ‘Dempsey’ leaf WT (left), leaf-derived callus tissue (middle), callus-derived emerging shoot tissue (right). ( B ) Volcano plots depicting the DEGs of callus versus WT (left) and shoot versus WT (right) comparisons. ( C ) Principal component analysis (PCA) plot of the TMM-normalized counts of the RNA-seq samples. ( D ) Correlation matrix plot (Corrplot) showing Pearson’s correlation efficient of RNA-seq samples. The filled fraction of the circle in each pie charts (upper) corresponds to the Pearson’s correlation coefficient (lower). Blue and red colors denote positive and negative correlations, respectively. ( E ) A heatmap of DEGs, which are grouped by K-means clustering into six clusters (colored bars) with numbers in brackets indicating the number of genes in each cluster. The X-axis represents the two biological replicates of RNA-seq samples taken from the three tissue types. The Y-axis represents individual gene expression levels, visualizing the variations in gene expression across tissue types and from the three tissue types. The Y-axis represents individual gene expression levels, visualizing the variations in gene expression across tissue types and samples. ( F ) Log 2 -transformed expression levels of genes in each K-means cluster. The X-axis represents the two biological replicates of RNA-seq samples taken from the three tissue types. The Y-axis represents the mean-centered log 2 expression level of the genes. Each graph is marked by a line representing the mean log 2 expression level in the color assigned to each cluster in panel E.

Article Snippet: Total RNA was isolated from ‘Dempsey’ leaf, callus, and shoot tissue using Tri-RNA Reagent (Favorgen, FATRR 001), and RNA concentrations were measured using a NanoDrop 2000 spectrophotometer (Thermo Fisher Scientific, Waltham, MA, USA; ND-2000).

Techniques: RNA Sequencing Assay, Derivative Assay, Expressing, Transformation Assay

Journal: BMC Plant Biology

Article Title: Elucidating the callus-to-shoot-forming mechanism in Capsicum annuum ‘Dempsey’ through comparative transcriptome analyses

doi: 10.1186/s12870-024-05033-4

Figure Lengend Snippet: Phytohormone-associated genes belonging to callus-specific Cluster 1 (red), shoot-specific Cluster 3 (aqua), and the cluster representing both callus and shoot tissue, Cluster 5 (pink), in C. annuum ‘Dempsey’. ( A ) a polar plot of phytohormone-related genes in each K-means cluster, with the eight phytohormones represented by each pole (see the light blue box for phytohormone abbreviations); ( B ) a stacked bar plot showing the gene numbers in each K-means cluster (colors) for each phytohormone (X-axis); ( C ) a proportional stacked bar plot of the genes in each K-means cluster (colors) for each phytohormone (X-axis); ( D ) heatmap of comparing the transcriptomes of the five species in callus tissue; ( E ) heatmap of comparing the transcriptomes of the five species in callus tissue. RNA-seq data were analyzed to identify phytohormone-related DEGs in each cluster with expression of C. annuum ‘Dempsey’ (red), A. thaliana (green), P. axillaris (blue), P. exserta (purple), and P. integrifolia (brown). The color scale bar of heat intensity indicates the log 2 -transformed fold change (log 2 |FC|) in expression (the grey box on the heatmap indicates no recorded expression). Red arrowheads indicate highly upregulated genes (log 2 |FC| > 2). The black arrowhead indicates the most upregulated gene (a log 2 |FC| of 1.5–2) for shoot formation (Cluster 3). The black boxes to the left of the heatmaps indicate the phytohormone(s) related to each gene

Article Snippet: Total RNA was isolated from ‘Dempsey’ leaf, callus, and shoot tissue using Tri-RNA Reagent (Favorgen, FATRR 001), and RNA concentrations were measured using a NanoDrop 2000 spectrophotometer (Thermo Fisher Scientific, Waltham, MA, USA; ND-2000).

Techniques: RNA Sequencing Assay, Expressing, Transformation Assay

Journal: ACS synthetic biology

Article Title: Efficient Reassignment of a Frequent Serine Codon in Wild-Type Escherichia coli

doi: 10.1021/acssynbio.5b00197

Figure Lengend Snippet: Northern blot analyses of intracellular aminoacylation of tRNAPyl isoacceptors. Total RNA was obtained from E. coli cells grown in the presence or absence of 3-I-Phe (1 mM) expressing IFRS and (A) either one or five gene copies of tRNAPylACU or (B) one gene copy of tRNAPylCAG, from the pCAM plasmid. Total RNA was suspended in either 10 mM sodium acetate, pH 4.5 (acylated), or 200 mM Tris, pH 8.0 (deacylated, OH−). Positions of acylated and deacylated tRNAPyl isoacceptors are indicated.

Article Snippet: Biochemical Characterization of the IFRS/tRNA Pyl Pair The aminoacylation assays were carried out for 60 min at 37 °C in aminoacylation buffer containing 100 mM Tris-HCl (pH 7.0), 10 mM MgCl 2 , 50 mM potassium chloride, and 5 mM DTT, with 1 mM 3-I-Phe (ChemImpex), 2 mM ATP, 500 nM IFRS, and 5 µM 32 P 3′-end-labeled transcript.

Techniques: Northern Blot, Expressing, Plasmid Preparation