Figure legends:
Figure 1. Subcellular localization of TaA6b Hsf. The ORF of TaA6bwas cloned in CaMV35S::TaA6b-GFP vector in which GFP was fused at the C-terminus. The TaA6b-GFP fusion protein and GFP control were transiently expressed in N. benthamiana leaves and observed by fluorescence microscopy. The left column is the green fluorescence channel, the center column is the DIC and the right column is the merge of the two images.
Figure 2. Effect of constant heat stress on transgenic lines (Line 2-1, Line 2-2 and Line 6-2) and wild type control plants (WT) in greenhouse with constant temperature at 35 ºC. Seeds were germinated in 2 inch nursery pots and kept in temperature controlled greenhouse from sowing and pictures were taken at specified time.
Figure 3.A. Detection of reactive oxygen species by NBT staining method in the leaves of barley plants from wild type (WT) and transgenic (AP2-2 and AP6-2) plants over-expressing wheat HSF (TaHsfA6b ) under control and heat stress conditions.
B. The activity of three different ROS scavenging enzymes viz ascorbate peroxidase (APX), catalase and super oxide dismutase (SOD) in wild type (WT) and transgenic (AP2-2 and AP6-2) plants under heat stress conditions. Error bars SD (n=5), P values **= <0.05 and ***= <0.001
Figure 4.A. Principle component analysis of RNA sequencing in WT (RCWT) and transgenic plants of two lines AP 2-1 and AP 2-2, showing grouping of biological replicates of WT and transgenic lines.
B. Volcano plot showing comparison of differentially expressed genes (DEGs) in wild type and transgenic barley plants. NS- Not significant , Log2 fold change- transcripts with fold change of >2 but non significant p-value, P-Transcript with p value<0.05, log2FC& P differentially expressed transcripts with log2FC >2 and p-value < 0.05.
Figure 5. qRT-PCR expression analysis of some upregulated genes in RNA-seq data (Supplementary table S2). Y-axis represent relative fold change calculated by 2^-ddct method.
Figure 6. Analysis of transgenic and WT plants under simulated drought stress treatment.A. 4 weeks-old wild type and transgenic plants of overexpression line AP2-2, under limited water condition.
B. Relative water content of leaves of wild type and transgenic plants of line AP2-2 kept under water limitation condition.