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Figure 9 in Evaluation of the Chilli veinal mottle virus CP gene expressing transgenic Nicotiana benthamiana plants for disease resistance against the virus

Figure 9. ELISA plate readings (O.D at 405nm) of leaf samples of transgenic lines (T1-T8) and control plants after 15 dpi.

Figure 7 in Evaluation of the Chilli veinal mottle virus CP gene expressing transgenic Nicotiana benthamiana plants for disease resistance against the virus

Figure 7. PCR Products of the hpt gene from T0 transgenic plants. Lane 1-9 are transgenic. Lane 10, +ve control. Lane 11, control (untransformed) plant.

Figure 6. DNA bands from T0 in Evaluation of the Chilli veinal mottle virus CP gene expressing transgenic Nicotiana benthamiana plants for disease resistance against the virus

Figure 6. DNA bands from T0 transgenic and Agro-infilterated plants. Lane 1-10, transgenic plants. Lane 11-13, agro-infilterated plants.

Figure 3 in Evaluation of the Chilli veinal mottle virus CP gene expressing transgenic Nicotiana benthamiana plants for disease resistance against the virus

Figure 3. Symptoms development on propagative host plants after mechanical inoculation with ChiVMV isolate ATIPK. (a) N. tabacum showing mosaic, mottling and vein clearing (b) C. annum (cv. Loungi) displays the symptoms of mottling, mosaic, leaf deformation and vein clearing.

Figure 2 in Evaluation of the Chilli veinal mottle virus CP gene expressing transgenic Nicotiana benthamiana plants for disease resistance against the virus

Figure 2. Symptoms of ChiVMV on chilli leaves collected from Islamabad. (a) Shows mottling and severe vein clearing and distortion. (b) Shows reduced leaf size with mottling and distortion.

opencc-by-4.0Feb 2022View details →

zenodo36/100

Dataset of results for a copper switch for inducing CRISPR/Cas9-based transcriptional activation tightly regulates gene expression in Nicotiana benthamiana.

CRISPR-based programmable transcriptional activators (PTAs) are used in plants for rewiring gene networks. Better tuning of their activity in a time and dose-dependent manner should allow precise control of gene expression. Here, we report the optimization of a Copper Inducible system called CI-switch for conditional gene activation in Nicotiana benthamiana. In the presence of copper, the copper-responsive factor CUP2 undergoes a conformational change and binds a DNA motif named copper-binding site (CBS). In this study, we tested several activation domains fused to CUP2 and found that the non-viral Gal4 domain results in strong activation of a reporter gene equipped with a minimal promoter, offering advantages over previous designs. To connect copper regulation with downstream programmable elements, several copper-dependent configurations of the strong dCasEV2.1 PTA were assayed, aiming at maximizing activation range, while minimizing undesired background expression. The best configuration involved a dual copper regulation of the two protein components of the PTA, namely dCas9:EDLL and MS2:VPR, and a constitutive RNA pol III-driven expression of the third component, a guide RNA with anchoring sites for the MS2 RNA-binding domain. With these optimizations, the CI/dCasEV2.1 system resulted in copper-dependent activation rates of 2,600-fold and 245-fold for the endogenous N. benthamiana DFR and PAL2 genes, respectively, with negligible expression in the absence of the trigger. The tight regulation of copper over CI/dCasEV2.1 makes this system ideal for the conditional production of plant-derived metabolites and recombinant proteins in the field.

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Activity-based proteomics reveals nine target proteases for the recombinant protein-stabilizing inhibitor SlCYS8 in Nicotiana benthamiana

Complete dataset (MS Label-free quantification) for the publication 'Activity-based proteomics reveals nine target proteases for the recombinant protein-stabilizing inhibitor SlCYS8 in Nicotiana benthamiana' DOI: 10.1111/pbi.13092

opencc-by-4.0Feb 2019View details →

opencc-by-4.0Jan 2022View details →

Nicotiana benthamiana and virus genomic references

Nicotiana benthamiana curated and modified genomic references. TSWV viral references.

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.

opencc-zeroOct 2020View details →

dryad28/100

Nicotiana benthamiana as a transient expression host to produce auxin analogues: Pisum sativum seed transcriptomic data

Plant secondary metabolites have applications for the food, biofuel, and pharmaceutical industries. Recent advances in pathway elucidation and host expression systems now allow metabolic engineering of plant metabolic pathways to produce "new-to-nature" derivatives with novel biological activities, thereby amplifying the range of industrial uses for plant metabolites. Here we use a transient expression system in the model plant Nicotiana benthamiana to reconstitute the two-step plant-derived biosynthetic pathway for auxin (indole acetic acid) to achieve accumulation up to 500 ng/g fresh mass (FM). By expressing these plant-derived enzymes in combination with either bacterial halogenases and alternative substrates, we can produce both natural and new-to-nature halogenated auxin derivatives up to 990 ng/g FM. Proteins from the auxin synthesis pathway, tryptophan aminotransferases (TARs) and flavin-dependent monooxygenases (YUCs), could be transiently expressed in combination with four separate bacterial halogenases to generate halogenated auxin derivatives. Brominated auxin derivatives could also be observed after infiltration of the transfected N. benthamiana with potassium bromide and the halogenases. Finally, the production of additional auxin derivatives could also be achieved by co-infiltration of TAR and YUC genes with various tryptophan analogues. Given the emerging importance of transient expression in N. benthamiana for industrial scale protein and product expression, this work provides insight into the capacity of N. benthamiana to interface bacterial genes and synthetic substrates to produce novel halogenated metabolites.

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Figure 10 in Evaluation of the Chilli veinal mottle virus CP gene expressing transgenic Nicotiana benthamiana plants for disease resistance against the virus

Figure 10. Symptoms development in transgenic plants after challenging with ChiVMV isolate ATIPK at15dpi. A) Resistance transgenic lines. B) Moderately resistance transgenic lines. C) Moderately susceptible transgenic lines.

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Figure 8 in Evaluation of the Chilli veinal mottle virus CP gene expressing transgenic Nicotiana benthamiana plants for disease resistance against the virus

Figure 8. PCR products from transgenic plants. (a) Lane 1-3, represent PCR product from agro-infilterated plants. Lane 4, control plant. (b) Lane 1-9, RT- PCR product from transgenic plants represent the presence of CP gene transcription. Lane 10, represents untransformed control sample.

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Figure 5 in Evaluation of the Chilli veinal mottle virus CP gene expressing transgenic Nicotiana benthamiana plants for disease resistance against the virus

Figure 5. Maximum likelihood tree of ChiVMV isolate ATIPK on nucleotide sequence of CP gene with 22 ChiVMV isolates from the world. The bootstrap analysis was conducted in 1000 replications. ZYMV (AB127936) sequence is used as out-group.