Gene expression raw count by RNA sequence

Two maize inbred lines with contrasting salt tolerance were used in the present study, of which L2010-3 is a salt-tolerant line and BML1234 is a salt-sensitive line. The seeds from the two lines were surface-sterilized in 10% (v/v) H2O2 for 15 min, rinsed with distilled water and then germinated on filter paper saturated with distilled water. Uniform seedlings with two leaves were transplanted into aerated 1 × Hoagland’s nutrient solution + Na150 for salt stress . The seedlings were then cultured in a growth room with a relative humidity of 70% (14/10 h, day/night) at 26℃. At 0 h (control), 6 h (T-6), 18 h (T-18), and 36 h (T-36) under salt treatment, the mixed roots from three seedlings of each line were individually collected as the samples for transcriptome sequencing, with two biological replicates.<br>According to the manufacturer’s instructions, total RNA of each sample was isolated using the HiPure Plant RNA Maxi Kit purchased from Magen company (http://www.magentec.com.cn). RNA library construction and transcriptome sequencing were performed by the Illumina sequencing platform.<br>Raw sequencing reads were filtered to remove low-quality reads, adaptor-polluted reads, and reads containing Poly-N, which generated clean reads by fastp (Chen et al. 2018). The clean reads were subsequently mapped to the maize reference genome (ZmB73 RefGen_V4) (http://www.gramene.org) via Hisat2. Gene expression levels were determined by Htseq software and were normalized to transcripts per million (TPM) by consumer script.