*4.5. Identification of Cis-Acting Elements*

A 2000 bp promoter region upstream of the MsHSF transcriptional start site was extracted from the *M. sativa* genome (http://www.genoscope.cns.fr/brassicanapus/) (accessed on 25 August 2022) and submitted to PlantCARE (http://bioinformatics.psb.ugent) (accessed on 25 August 2022) to identify the three types of regulatory cis-acting elements and finally visualize the results using TBtools.

#### *4.6. Analysis of Tissue-Specific Expression and Abiotic Stress Transcriptome Data*

Relevant transcriptome data were downloaded from the public NCBI database to investigate the expression patterns of *MsHSF* genes in different tissues and under different biotic stresses. *M. sativa* RNA-Seq data for different tissues (pre-elongated stems, elongated stems, flowers, leaves, roots, and rhizomes) can be downloaded from accession number SRP055547. The NCBI Short Read Long Archive database contains abiotic stressrelated transcriptome data (cold, at SRR7091780-SRR7091794; drought, salt, and ABA, at SRR7160313-SRR7160357). The raw data was filtered and the SRA files were converted to FASTQ files by the SRA to Fastq program of TBtools. Gene expression levels were calculated by fragment number per kilobase per million mapped reads (FPKM values). Finally, the Hetmap program of TBtools was imported to generate the associated heat map.
