Project description:Purpose: Investigate genes associated with Phn7.1, a major QTL influencing partial resistance to the soil-borne pathogen Phytophthora nicotianae in tobacco. Methods: Resistant and susceptible tobacco near isogenic lines with and without Phn7.1 QTL were subjected to the inoculation with Phytophthora nicotianae suspension and suspension buffer without pathogen as control followed by sample collection at 42 hour past inoculation for RNA-seq analysis. Results: Revealed gene expression profiles associated disease resistance and susceptiblilty.
Project description:For decades the tobacco plant has served as a model organism in plant biology to answer fundamental biological questions in the areas of plant development, physiology, and genetics. Due to the lack of sufficient coverage of genomic sequences, however, none of the expressed sequence tag (EST)-based chips developed to date cover gene expression from the whole genome. The availability of Tobacco Genome Initiative (TGI) sequences provides a useful resource to build a whole genome exon array, even if the assembled sequences are highly fragmented. Here, the design of a Tobacco Exon Array is reported and an application to improve the understanding of genes regulated by cadmium (Cd) in tobacco is described. From the analysis and annotation of the 1,271,256 Nicotiana tabacum fasta and quality files from methyl filtered genomic survey sequences (GSS) obtained from the TGI and ~56,000 ESTs available in public databases, an exon array with 272,342 probesets was designed (four probes per exon) and tested on two selected tobacco varieties. Two tobacco varieties out of 45 accumulating low and high cadmium in leaf were identified based on the GGE biplot analysis, which is analysis of the genotype main effect (G) plus analysis of the genotype by environment interaction (GE) of eight field trials (four fields over two years) showing reproducibility across the trials. The selected varieties were grown under greenhouse conditions in two different soils and subjected to exon array analyses using root and leaf tissue to understand the genetic make-up of the Cd accumulation. An Affymetrix Exon Array was developed to cover a large (~90%) proportion of the tobacco gene space. The Tobacco Exon Array will be available for research use through the Affymetrix array catalogue. As a proof of the exon array usability, we have demonstrated that the Tobacco Exon Array is a valuable tool for studying Cd accumulation in tobacco leaves. Data from field and greenhouse experiments supported by gene expression studies strongly suggested that the difference in leaf Cd accumulation between the two specific tobacco cultivars is dependent solely on genetic factors and genetic variability rather than on the environment. 22 samples were used with 3 different experimental factors: [1] 2 different tissues (leaves and roots/lateral), [2] 2 different mutations (V5 and V21), and [3] 2 different stimuli (Soil 1 and Soil 2). 2-3 biological replicates were used. One sample did not pass QC and is not included in this submission.
Project description:The soil-borne bacterial pathogen Ralstonia solanacearum invades a broad range of plants through roots, resulting in wilting of the plant, but no effective protection against this disease has been developed. Two R. solanacearum resistance-inducing compounds were biochemically isolated from tobacco and identified as sclareol and cis-abienol, diterpenes. When exogenously applied to their roots, these diterpenes induced resistance to R. solanacearum in tobacco, tomato, and Arabidopsis plants without exhibiting any antimicrobial activity. Structure-activity correlation analysis of sclareol-related compounds revealed that the hydroxyl group at the eighth carbon position is responsible for the activity for inducing resistance. Microarray analysis identified many sclareol-responsive Arabidopsis genes, such as those encoding or with role in ABC transporters, biosynthesis and signaling of defense-related signal molecules, and mitogen-activated protein kinase (MAPK) cascades. Sclareol-induced R. solanacearum resistance was partially compromised in Arabidopsis mutants defective in the ABC transporter AtPDR12, the MAPK MPK3, and ethylene and abscisic acid signaling pathways. Transgenic tobacco plants in which NtPDR1, a tobacco homolog of AtPDR12, was silenced exhibited also reduced resistance. These results suggest that multiple host factors are involved in resistance to R. solanacearum induced by sclareol and its related compounds and that these compounds can be used to protect crops from bacterial wilt disease.
Project description:Spider silk, especially dragline silk from Trichonephila clavipes, is an excellent natural material with remarkable mechanical properties. In this study, we generated nucleus- and plastid-encoded expression of a partial sequence of one of the components of dragline silk, major ampullate spidroin-1 (MaSp1), in tobacco (MaSp1-tobacco), MaSp1nuc and MaSp1pla, respectively. The partial sequence of MaSp1 encodes six repetitive glycine-rich and polyalanine tandem domains, and we evaluated the effect of its expression on plant mechanical properties and physiology. While MaSp1 expression showed no effect on leaf mechanical properties, it conferred drought tolerance. MaSp1-tobacco demonstrated an increased growth on PEG-infused growth medium and higher drought recovery in soil compared to the non-transgenic control (WT). Transcriptome analyses of drought-stressed MaSp1-tobacco revealed an upregulation of genes involved in stress response, antioxidant activity, and phenylpropanoid biosynthesis, with a higher effect of drought on the MaSp1pla transcriptome. Genes encoding peroxidase, stress and channel proteins, and cellular metabolism and homeostasis were consistently upregulated in these lines, which also displayed a higher total antioxidant status (TAS) compared to WT, and a higher TAS in MaSp1pla compared to MaSp1nuc. Under normal watering conditions, the ABA content and the TAS in MaSp1-tobacco were significantly higher than in the WT. Our results show that nucleus- and plastid-encoded expression of MaSp1 in tobacco increased the endogenous ABA levels, which led to the formation of plant stress memory that caused overall adjustments in multiple drought tolerance mechanisms and hence, conferred drought tolerance.
Project description:Background :Nitrogen (N) supply directly impacts growth and quality in flue-cured tobacco. To decipher molecular responses to N gradients, we integrated transcriptomics and weighted gene co-expression network analysis (WGCNA) on leaves from four N treatments: 0 (inherent soil fertility), 60 (low), 105 (standard), and 150 kg/hm² (high). Results :Phenotypic analysis revealed dose-dependent increases in leaf nitrogen content with higher N application, accompanied by excessive vegetative growth and delayed maturity at 150 kg/hm². Transcriptome sequencing identified 47,216 genes, with differentially expressed genes (DEGs) increasing linearly with N levels (1,458–2,147 DEGs). Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment highlighted nitrogen metabolism pathways, yielding 14 DEGs (11 in assimilation, 3 in transport). Weighted gene co-expression network analysis (WGCNA) uncovered two modules (lightcyan1 and black) strongly associated with N responses, harboring transcription factors NtERF11 (AP2/ERF), NtWRKY3 (WRKY), and NtSRM1 (MYB). Sub-network analysis within these modules identified five hub genes: NtGLN1-1, two uncharacterized genes, NtDFC , and NtGDSL. NtGDSL may enhance nitrogen use efficiency (NUE) through stress-responsive mechanisms, while NtDFC could integrate N signaling with developmental processes. These findings provide novel insights into N regulatory networks in flue-cured tobacco. Conclusions :This study reveals the effects of nitrogen application rates on flue-cured tobacco growth and gene expression. By identifying key transcription factors and genes regulating nitrogen metabolism, it provides a theoretical basis for dissecting nitrogen regulatory mechanisms, optimizing fertilization strategies, and improving nitrogen use efficiency in tobacco production.