*4.5. Plant Material Preparation and Transcriptomic Data Analysis*

*C. songorica* seeds were sown in vermiculite medium supplied with 1/4 diluted Hoagland's nutrient solution, pH 5.8. Growth chamber conditions were set at 75–80% relative humidity, 30/28 ◦C (day/night), and 16/8 h (day/night) light at 200 mmol photons m−<sup>2</sup> s −1 . One-month old seedlings were treated with 40 ◦C or with 100 µM ABA. Root and shoot tissue samples were collected at 0 and 24 h post the treatment and kept at −80 ◦C till RNA extraction. Three root and shoot samples were collected from each treatment. Total RNA was isolated from the samples using the Shengong RNA isolation kit as instructed (Shengong Ltd., Shanghai, China). RNA pools were constructed following Illumina sequencing guidelines and then sequenced conferring to RNA-seq procedure. In total 24 million 250-bp raw reads were produced from the 12 samples. To eliminate adapter sequences from raw reads, the FASTX version 0.0.13 toolkit (http://hannonlab.cshl.edu/fastxtoolkit/) was used. Additionally, the FastQC server tool (http://www.bioinformatics.babraham.ac.uk/projects/fastqc/) was utilized to assess the quality of sequences. The resulting clean reads were aligned with the *C. songorica* genome by means of Tophat v.2.0.10 server (http://tophat.cbcb.umd.edu/) [67] and the produced alignment files were used as Cufflinks inputs to create transcriptome assemblies [68]. The *C. songorica* gene expression levels were estimated based on fragments per kilobase of exon model per million mapped reads (FPKM) for root and shoot tissues. Following this, a random sampling model built on read count for each individual gene was applied to determine differentially expressed [69]. Gene expression levels

were normalized with Pearson coefficients to generate hierarchical clustering with average linkage using HemI toolkit [70].
