3.2.3 Impact of temperature on scent related genes
To check whether elevated air temperature regime affects the end products of scent pathway, the expression levels of gene-encoding enzymes responsible for the formation of volatile compounds were evaluated. Based on earlier information about the higher transcript accumulation levels in J. auriculatum flowers under in situ condition, extractions of total RNAs were made from flowers ofJ. auriculatum plant grown at four different air temperatures as defined earlier with at least two/three biological replication.
It was observed that expression levels of Ja PAL, JaBEATwere high in flowers of J. auriculatum grown at 25°C; the accumulation of JaMTS transcript was higher at both 25°C and 30°C while the accumulation of JaPAR was highest at 30°C (Figure 7a, b ). The relative expression levels of JaBEATand JaMTS monitored via RT-qPCR correlated well with semi-quantitative RT-PCR results (Figure 7c ). Elevated accumulations of JaHMGS and JaHMGR transcripts were observed at both 25°C and 30°C. Successive upliftmenst of Jaβ-Gluand JaMYB transcripts were observed in flowers of J. auriculatum grown at 25°C followed by 30°C air temperature. The expression levels of all these genes were much lower at air temperatures of 20°C and 35°C.
3.2.4 Influence of temperature onaccumulation of non-volatile metabolite compounds
Different groups of non-volatile metabolites were identified from polar floral extracts. Mature buds fromJ. auriculatum plants growing under different temperature regime were collected and subsequently freeze-dried aiming at obtaining the maximum recovery of non-volatile metabolites. Compounds comprising mainly of primary metabolites such as, sugars, organic acids, fatty acids and other specialized metabolites (mainly phenolic acids) which are non-volatile in nature were detected as derivatized metabolites from lyophilized flowers. These metabolites were compared from plants grown at different temperatures to monitor any significant perturbations that might have occurred in the primary metabolism. In total, 37 metabolites were identified from GC-MS chromatogram (Figure S2, Table S2) . The contents of these metabolites are represented in the form of a heatmap illustration with colour gradation to signify the changes under different air temperature regime (Figure 8a ). An overall reduction in the contents of non-volatile metabolites was observed at 20°C, the lower range border air temperature set out for this study. Enhanced accumulations of most of the sugar metabolites viz. fructose, glucose, lactose, lactulose, gentiobiose and mannobiose were observed in flowers of J. auriculatum grown at 25°C subsequently followed by 30°C. Enhanced abundance of benzenoid precursors such as, coutaric acid, caffeic acid and shikimic acid were found in flowers of J. auriculatum plant grown at 25°C; the content of 4-hydroxy benzyl alcohol was also found to be highest in flowers of 30°C temperature grown plants followed by 25°C. The contents of most of these detected phenolics were however low both at 20°C and 35°C. Further the relative abundance of mevalolactone, the lactone form of mevalonic acid (which serves as precursor for biosynthesis of sesquiterpenes), was found to be highest in flowers of J. auriculatum plant grown at 30°C. The abundance of major organic acids was highest in plants growing at 30°C. At 35°C, diversion of metabolites from volatile pathway towards sugar alcohols viz. fucitol, glucitol and myo-inositol was observed. PCA was carried out upon using the metabolites as variables (Figure 8b, c ). A clear segregation of all temperature treated samples was observed with well clustered individual sample replicate. According to score plot, treatments at 25°C and 30°C were closer on the PC1 axis. However, the plants grown at 20°C and 35°C, (the two border-range air temperatures set out for conducting these experiments) had some proximity as observed in terms of PC2.