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, Notable reduction in planta multiplication of R. solanacearum was observed under SD-combined stress compared to SD-pathogen stress (Supplementary Figure S2B). However, in planta bacterial count was unchanged under LD-combined stress compared to LD-pathogen stress treatments (Supplementary Figure S2C). Transcriptomic alterations in chickpea plants challenged with combined drought and R. solanacearum stress (SD-and LDcombined) and individual drought (SD-and LD-drought) and R. solanacearum stress (SD-and LD-pathogen) were studied by microarray analysis. Microarray data was submitted to Gene Expression Omnibus (GEO# GSE89228). The DEGs, FC-35%) and LD drought (FC-30%) showed 73 and 52% RWC, respectively, compared to 86% RWC in control plants after 10 and 15 days of drought treatment, respectively. R, p.3827

, Similarly, under SD-drought and LD-drought stress treatments, genes involved in signaling, biosynthesis of abscisic acid (ABA) and osmo-protectants namely the genes encoding for LATE EMBRYOGENESIS ABUNDANT 5 (LEA5), LOW-TEMPERATURE-INDUCED 65 KDA PROTEIN were upregulated. Concordant with these observations, genes involved in both defense responses and abiotic stress tolerance (genes encoding for LEA and RESPIRATORY BURST OXIDASE HOMOLOG B) were differentially expressed in SD-combined and LD-combined stresses, which conforms to the nature of the stressors (Supplementary Figure S5 and File S1). The majority of top most up-regulated genes were belonging to stress responsive and cell wall modification categories under combined stress (Supplementary Figure S5 and File S1). The number of up-and down-regulated genes under each stress condition is shown in Figure 1. The maximum numbers of DEGs were found in LD drought stress (1426 genes). Comparison of DEGs among SD stress treatments showed 821 genes (31.8%) out of 1011 genes to be uniquely up-regulated in response to SD-combined stress whereas, SD-combined stress treatment had 129 and 58 DEGs in common with the SD-pathogen and SD-drought stress, respectively (Figure 1B). Similarly, 1039 genes (31.5%) out of 1287 total DEGs were uniquely expressed under LD combined stress (Figure 1C). LD-combined stress and LD-drought stress had 102 DEGs in common, Under both SD-pathogen and LD-pathogen stress treatments, many genes involved in defense response (WRKY33, MAP KINASE 11, and DEFENSIN) were up-regulated

, As a result a substantial variation in stress responsive transcriptome under SD-and LD-stresses was observed. The number of DEGs was more in LD stresses over SD stresses. For instance, number of DEGs under LD-pathogen was 841 compared to 594 under SD-pathogen stress treatment. Similarly, LD-drought stress treatment, SD-combined and LD-combined stress transcripts (Figures 1D-F)

, LD-pathogen, LD-drought and LD-combined stress had 707, 1000, and 946 unique DEGs, respectively, as compared to 460, 778, and 709 unique DEGs in response to SD-pathogen, SDdrought and SD-combined stress, respectively. Moreover, very small percentages (10-15%) of genes were common between the respective SD and LD treatments (Figures 1D-F). The overlapping genes showed differential expression under different stresses. A few common genes between LD combined and LD drought stress (genes encoding for LEA5, E3 UBIQUI TIN-PROTEIN LIGASE, PP2C37, INOSITOL 3 PHOSPHATASE SYNTHASE LIKE, and MATE EFFULUX FAMILY PROTEIN 5) had higher expression in LD-combined stress as compared to LD-drought stress, DEGs under SD-combined stress. Each stress transcriptome had more number of unique DEGs and less number of common DEGs

, Color scale shows gene expression range where color bar in red and blue represents up-and down-regulated genes, respectively. Details of the genes shown in heat maps are available in Supplementary File S5. RESISTANCE 3, ZINC FINGER PROTEIN, DOF ZINC FINGER PROTEIN DOF1.1, SER/THR-PROTEIN KINASE EDR1, (STPKEDR1) RESPIRATORY BURST OXIDASE HOMOLOG B (RBOHB), TIR CLASS DISEASE RESISTANCE, and NITRATE REDUCTASE under SD-pathogen stress. LD-pathogen stress showed up-regulation of defense related genes encoding RETI CULIN OXIDASE LIKE PROTEIN, DISEASE RESISTANCERESPONSIVE (dirigent-like protein), GLUTAMINE AMIDO TRANSFERASE, PR, BOTRYTIS SUSCEPTIBLE 1, WRKY70, and RPP13. SD-combined stressed plants exhibited high upregulation of genes encoding RETICULIN OXIDASE LIKE PROTEIN (also called BERBERINE BRIDGE ENZYME), FIGURE 2 | Expression profile of DEGs common between combined and individual stresses. The DEGs with more than one fold expression and p < 0.05 under SD treatments (SD-pathogen, SD-drought, SD-combined stress) and LD treatments (LD-pathogen, LD-drought, 1991.

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