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Fig. 6. A in Comparative analysis of cyclotide-producing plant cell suspensions presents opportunities for cyclotide plant molecular farming

Fig. 6. A Structural equivalence of cyO2 derived and purified from plant suspension cells or whole plant tissue as determined by NMR spectroscopy. B Cytotoxicity of cyclotides cyO2 and kB1 against Sf9 cells.

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Fig. 4 in Comparative analysis of cyclotide-producing plant cell suspensions presents opportunities for cyclotide plant molecular farming

Fig. 4. Inducibility of cyclotide accumulation in O. affinis suspension cells. A Cyclotide accumulation profiles in O. affinis suspension cells exposed to biotic (methyl jasmonate, MeJA) and abiotic (NaCl) signals. Replicate (n =3) cultures were assayed 1, 4 and 7 days after addition of inducer. Treatments highlighted by an asterisk are significantly different (P <0.05). Cell morphology of B day 1 cells in 3 mM NaCl, C day 7 cells in 3 mM NaCl and D day 7 cells in 50 mM NaCl.

Fig. 2 in Comparative transcriptome analysis infers bulb derived in vitro cultures as a promising source for sipeimine biosynthesis in Fritillaria cirrhosa D. Don (Liliaceae, syn. Fritillaria roylei Hook.) - High value Himalayan medicinal herb

Fig. 2. (A–E) Differential gene expression analysis in comparative F. roylei transcriptome: (A) Heat-map showing differential gene expression in the bulb (PKW) vs callus (PK2); bulb (PKW) vs in vitro regenerated plantlets (PK1) and callus (PK2) vs in vitro regenerated plantlets (PK1); (B) Venn diagram represents the differential gene expression in PKW vs PK1; PKW vs PK2; PK2 vs PK1, (C–E) Volcano plots represents the differential gene expression in PKW vs PK1; PKW vs PK2; PK2 vs PK1 as colour description image, where p-value & log2 fold-change in red colour represents genes with log2 fold-change cut off 2 and p-value <=0.05; p-value in blue colour represents genes with no cut off on log2 fold-change and p-value <=0.05. Whereas, log2 fold-change in green colour represents genes with fold-change cut off 2 but no p-value cut off and non-significant (NS) in grey colour represents genes with no filter on log2 fold-change and p-value, respectively. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig. 1 in Comparative transcriptome analysis infers bulb derived in vitro cultures as a promising source for sipeimine biosynthesis in Fritillaria cirrhosa D. Don (Liliaceae, syn. Fritillaria roylei Hook.) - High value Himalayan medicinal herb

Fig. 1. (A–F) Functional annotations and unigenes classification of comparative F. roylei transcriptome: (A) Unigenes annotation with top 15 different plant species; (B) Top 5 pathway representation as per Kyoto Encyclopedia of Genes and Genomes; (C) Gene Ontology classification under the cellular component, molecular function, and biological process categories; (D) COG (Cluster of Orthologous Groups of proteins) classification into nine different categories; (E) Unigenes classification into major transcription factor families; (F) Gene family and sub-family classification using TAIR database.

Fig. 4 in Comparative transcriptome analysis infers bulb derived in vitro cultures as a promising source for sipeimine biosynthesis in Fritillaria cirrhosa D. Don (Liliaceae, syn. Fritillaria roylei Hook.) - High value Himalayan medicinal herb

Fig. 4. Comparative expression pattern validation for the sipeimine biosynthetic pathway genes as obtained from RNA-Seq data and qRT-PCR.

Fig. 3 in Comparative transcriptome analysis infers bulb derived in vitro cultures as a promising source for sipeimine biosynthesis in Fritillaria cirrhosa D. Don (Liliaceae, syn. Fritillaria roylei Hook.) - High value Himalayan medicinal herb

Fig. 3. Proposed sipeimine biosynthetic pathways in F. roylei. Heat-map showing gene expression in the bulb, callus, and regenerated plantlets. The genes were mapped using TPM (transcripts per kilobase million) values and colour-coded by increasing relative expression. The broken dotted arrow represents putative terminal biosynthesis steps. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig. 6 in Comparative transcriptome analysis reveals the regulatory effects of acetylcholine on salt tolerance of Nicotiana benthamiana

Fig. 6. Differentially expressed genes related to the plant hormone signal transduction pathway in the comparison (NaCl vs NaCl + ACh). (A) diagram of auxin, gibberellin, brassinosteroid and salicylic acid signalling transduction pathways; (B) information and expression patterns of differentially expressed genes involved in auxin, gibberellin, brassinosteroid and salicylic acid signalling transduction pathways. Red means upregulated expression of genes, and green means downregulated expression of genes. The number in each sample name represents the sample order. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig. 3 in Comparative transcriptome analysis reveals the regulatory effects of acetylcholine on salt tolerance of Nicotiana benthamiana

Fig. 3. Functional annotation of differentially expressed genes (DEGs) based on gene ontology categorization. The left Y-axis represents the significantly enriched GO terms (p <0.05) pathways. The Xaxis represents the percentage of DEGs belonging to the corresponding pathway. The sizes of bubbles represent the number of DEGs in the corresponding pathway, and the colours of the bubbles represent the enrichment p-value of the corresponding pathway. The left y-axis shows the Gene Ontology terms. Biological process, cellular component and molecular function are indicated by different colours. Only significantly enriched GO terms (p <0.05) are shown. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig. 8 in Comparative transcriptome analysis reveals the regulatory effects of acetylcholine on salt tolerance of Nicotiana benthamiana

Fig. 8. Quantitative real-time PCR (RT-qPCR) validation of selected differentially expressed genes detected in Nicotiana benthamiana leaves. The expression levels obtained by RT-qPCR are represented in black lines, RT-qPCR data showed the mean values from three replicates, and the error bars represent the SE of the means, while the corresponding expression data for RNA-seq are represented in the white histogram. CN, control; CN + ACh, 10 μM acetylcholine; NaCl, 150 mM NaCl stress; NaCl + ACh, 150 mM NaCl stress plus 10 μM acetylcholine.

Fig. 5 in Comparative transcriptome analysis reveals the regulatory effects of acetylcholine on salt tolerance of Nicotiana benthamiana

Fig. 5. Heatmap representing the differentially expressed genes (DEGs) involved in cell wall extensibility of Nicotiana benthamiana leaves as influenced by NaCl alone or in combination with acetylcholine treatment (NaCl + ACh). Red means upregulated expression of genes, and green means downregulated expression of genes. The number in each sample name represents the sample order. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig. 1 in Comparative transcriptome analysis reveals the regulatory effects of acetylcholine on salt tolerance of Nicotiana benthamiana

Fig. 1. Changes in reactive oxygen species accumulation and production and antioxidant enzyme activity in Nicotiana benthamiana leaves 5 days after treatment with acetylcholine (ACh) under salt stress. (a) leaves were stained with NBT and DAB, (b) Fv/Fm, (c) Superoxide content, (d) Hydrogen peroxide content, (e) Ascorbate peroxidase activity and (d) Catalase activity as influenced by salt stress alone or in combination with ACh treatment. Data are means of three replications ±SE. Means with the same lowercase letters are not significantly different at p <0.05, according to Duncan's multiple range test. CN: control; CN + ACh, 10 μM acetylcholine; NaCl, 150 mM NaCl stress; NaCl + ACh, 150 mM NaCl stress plus 10 μM acetylcholine.

Fig. 7 in Comparative transcriptome analysis reveals the regulatory effects of acetylcholine on salt tolerance of Nicotiana benthamiana

Fig. 7. Heatmap representing the differentially expressed genes involved in transcription factors extensibility of Nicotiana benthamiana leaves as influenced by NaCl alone or in combination with acetylcholine treatment (NaCl + ACh). Red means upregulated expression of genes, and green means downregulated expression of genes. The number in each sample name represents the sample order. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig. 4 in Comparative transcriptome analysis reveals the regulatory effects of acetylcholine on salt tolerance of Nicotiana benthamiana

Fig. 4. The Kyoto Encyclopedia of Genes and Genomes pathway enrichment scatter map (p <0.05). The X-axis (Rich Factor) represents the percentage of DEGs belonging to the corresponding pathway.

Fig. 1 in Comparative metabolomics analysis of the response to cold stress of resistant and susceptible Tibetan hulless barley (Hordeum distichon)

Fig. 1. HCA and PCA of metabolite profiles under different temperatures of different varieties. (A) Heatmap of the metabolites detected in the total samples. Red indicates high abundance, green indicates low abundance. Metabolites were divided into three main clusters (1, 2, 3). (B) Score plot of PCA of metabolite datasets of the different samples. Each point represents one metabolite profiling experiment. There were three biological repeats for each temperature. 1/2/3/4/5/6 represent the different temperatures: 24 °C/12 °C/ 5 °C/0 °C/–5 °C/–8 °C, respectively. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig. 5 in Comparative metabolomics analysis of the response to cold stress of resistant and susceptible Tibetan hulless barley (Hordeum distichon)

Fig. 5. Abundance of the metabolites enriched in glutathione metabolism during cold treatment. Fold changes in metabolite levels under the indicated temperatures in XL and ZQ are shown; red represents upregulation, blue represents downregulation. (For interpretation of the references to colour in this figure legend, the reader is referred to the Web version of this article.)

Fig. 4 in Comparative metabolomics analysis of the response to cold stress of resistant and susceptible Tibetan hulless barley (Hordeum distichon)

Fig. 4. Analysis of DAMs in XL and ZQ. (A) Venn diagram of DAMs between cold stress and control conditions (ZQ1) in ZQ. (B) Venn diagram of DAMs between cold stress and control conditions (XL1) in XL. (C, D) Top 10 upregulated and downregulated DAMs in XL (C) and ZQ (D) plants exposed to −8 °C or 24 °C.

Fig. 3 in Comparative metabolomics analysis of the response to cold stress of resistant and susceptible Tibetan hulless barley (Hordeum distichon)

Fig. 3. DAMs in XL and ZQ plants exposed to freezing stress compared to the control condition. Metabolites significantly altered in XL, but not in ZQ plants exposed to freezing stress compared to the control condition (24 °C).

Fig. 2. DAMs between XL in Comparative metabolomics analysis of the response to cold stress of resistant and susceptible Tibetan hulless barley (Hordeum distichon)

Fig. 2. DAMs between XL and ZQ. (A) Numbers of DAMs under cold stress (12 °C, 5 °C, 0 °C, −5 °C, −8 °C) or the control condition (24 °C) in XL and ZQ. 1/2/3/4/5/ 6 represent 24 °C/12 °C/5 °C/0 °C/–5 °C/–8 °C, respectively.(B) Upregulated and downregulated metabolites in XL and ZQ at different temperatures. 1/2/3/4/5/6 represent 24 °C/12 °C/5 °C/0 °C/–5 °C/–8 °C, respectively. (C, D) Metabolites that were significantly altered in XL exposed to cold stress (12 °C, 5 °C, 0 °C) compared to the control condition (24 °C), but not in ZQ.