title: RNA-Seq of Aphanamixis polystachya: control
design: Transcriptome analysis was carried out in the seedlings kept in 50 mM salicylic acid on the 5th day and two-month-old seedlings (control).  RNA was isolated from Shoot and Leaf from Control and Treated plants by RNAeasy plant mini kit combined with the CTAB method. Isolated RNA of samples control seedlings (shoot) and (leaves) and stressed seedlings (shoot) and (leaf) were pooled by equimolar concentration and subjected to library preparation.  The libraries were prepared with input total RNA (~1 g) using Illumina TruSeq Stranded Total RNA Library Preparation Kit as per the manufacturers protocol.  Briefly, total RNA was Ribo depleted using plant rRNA removal mix and rRNA removal beads, then subjected to purification, fragmentation and priming for cDNA synthesis. The Ribo depleted fragmented RNA was converted into the first-strand cDNA, followed by second-strand cDNA synthesis, A-tailing, adapter-index ligation and finally amplified by the recommended number of PCR cycles. Library quality and quantity check were performed using Agilent DNA High Sensitivity Assay Kit. The amplified libraries were analyzed on Bioanalyzer 2100 (Agilent Technologies) using High Sensitivity (HS) DNA chip as per manufacturer's instructions.  After obtaining the Qubit concentration for the library and the mean peak size from Bioanalyzer profile, libraries were loaded into the Illumina platform for cluster generation and sequencing.  Paired-End sequencing allowed the template fragments to be sequenced in both the forward and reverse directions. The library molecules bind to complementary adapter oligos on the paired-end flow cell. The adapters were designed to allow selective cleavage of the forward strands after re-synthesis of the reverse strand during sequencing. The copied reverse strand was then used to sequence from the opposite end of the fragment.  The sequenced raw data from Illumina BCL format were converted to Fastq format using bcl2fastq software (Illumina). These Fastq data were filtered using Trimmomaticver 0.36, and high-quality reads were produced. From this high-quality reads Denovo assembly was carried out by CLC bios workbench, and unigenes were generated.