biostudies-arrayexpressLUIS ROMEROOryza sativa Japonica GroupBismarkhttps://www.ebi.ac.uk/biostudies/studies/E-MTAB-15231To investigate the effect of H2S on DNA methylation of plants submitted to drought stress, we performed transcriptome sequencing analysis in four experimental treatments: rice watered plants (control sample, C), rice plants watered with NaHS solution, as sulfide donor (sulfide treatment, S), rice plants subjected to drought stress (drought, D) and rice plants pretreated with NaHS and subjected to drought stress (S_D). Thus, 25-day-old plants grown in soil under physiological conditions were divided into two batches with one batch irrigated with water and the other with NaHS for 10 days, replacing the NaHS solution every two days. After this period, each batch was subsequently divided into two new batches and subjected to water irrigation or drought for another 15 additional days. At the end of the full treatment, four replicates of each condition were obtained that were processed independentlybiostudies-arrayexpressSequencing - Quantified libraries are pooled and sequenced on an Illumina platform and paired-end reads were generated. Before analysis, the raw sequencing data underwent quality assessment using FastQC (fastqc_v.0.11.5) and processed through Trimmomatic (Trimmomatic-0.36) software. All the reads that passed all the filtering steps was counted as clean reads and used for subsequent analyses. Bismark software (versión 0.16.3) (Krueger and Andrews, 2011) was used to perform alignments of bisulfite-treated reads to the reference genome (ensemblplants_oryza_sativa_japonica_ group_irgsp_1_0_gca_001433935_1). The bisulfite conversion rate, as the proportion of methylated clean reads to the reference genome, was calculated. The methylation information of cytosine sites was extracted based on the principle of cytosine–thymine conversion. The unmethylated cytosine was converted to thymine, while the methylated cytosine remained unchanged. Cytosine sites were classified into CpG, CHG, and CHH contexts (where H stands for A, T, and C). The binomial distribution test was used to determine methylation status and cytosine sites. Genome-wide methylation levels were analyzed using a 10,000 bp window size and the sum of methylated and unmethylated read counts in each window calculated. The methylation level of each window or C site is defined as: ML = reads(mC) / reads(mC) + reads (C). Differentially methylated regions (DMRs) were identified using the DSS software. According to the distribution of DMRs through the genome, we defined the genes related to DMRs as genes whose gene body region or promoter region have an overlap with the DMR.Sample Treatment - Germinated seeds were sown in soil and grown for 25 days under physiological water regime. Then, plants were divided into two batches, one water-irrigated and other treated with50 μM NaHS (as hydrogen sulfide donor compound) for an additional 10 d. After this period, each batch was subsequently divided into two new batches and subjected to water irrigation or drought for another 15 d. At the end of the whole treatment, four different plant samples were obtained as follows: control (C, water-irrigated plants during all period), sulfide-treated (S, sulfide-treated plants without any drought stress), drought (D, drought-treated plants with no-sulfide treatment), and sulfide-pretreated plus drought (S_D, sulfide-pretreated and after drought subjected plants). Leaves from different plant samples were collected, frozen in liquid nitrogen and stored at -80 C for further analysis.Sample Collection - Germinated seeds were sown in soil under a photoperiod of 12 h of white light (100 μmol m–2 s–1) at 28 C and 12 h of darkness at 28 C, and grown for 25 days under physiological water regime. Then, plants were divided into two batches, one water-irrigated and other treated with50 μM NaHS (as hydrogen sulfide donor compound) for an additional 10 d. After this period, each batch was subsequently divided into two new batches and subjected to water irrigation or drought for another 15 d. At the end of the whole treatment, four different plant samples were obtained as follows: control (C, water-irrigated plants during all period), sulfide-treated (S, sulfide-treated plants without any drought stress), drought (D, drought-treated plants with no-sulfide treatment), and sulfide-pretreated plus drought (S_D, sulfide-pretreated and after drought subjected plants). Leaves from different plant samples were collected, frozen in liquid nitrogen and stored at -80 C for further analysis.Library Construction - Fragmented DNA of 200-400 bp was bisulfite treated to generated single strand DNA in which unmethylated cytosine were converted into uracil and methylated cytosine unchanged. Double strand DNA was synthesized, subjected to PCR amplification, and the library was ready after size selection.Nucleic Acid Extraction - DNA was extracted from leaves of two plant samples: control (C, water-irrigated plants during all period) and sulfide-treated (S, sulfide-treated plants without any drought stress) using the Qiagen DNeasy Plant Mini Kit. Four independent biological replicates of each sample were usedGrowth Protocol - Germinated seeds were sown in soil under a photoperiod of 12 h of white light (100 μmol m–2 s–1) at 28 C and 12 h of darkness at 28 C, and grown for 25 days under physiological water regime. Then, plants were divided into two batches, one water-irrigated and other treated with50 μM NaHS (as hydrogen sulfide donor compound) for an additional 10 d. After this period, each batch was subsequently divided into two new batches and subjected to water irrigation or drought for another 15 d.OrganizationMINSEQE ScoreAssays and DataProcessed DataMAGE-TAB FilesData Transformation - Differentially methylated regions (DMRs) were identified using the DSS software (Hao Feng HaoWu, 2014 ； Hao Wu ， 2015 ； Yongseok Park Hao Wu,2016 ) , The core of DSS is a new dispersion shrinkage method for estimating the dispersion parameter from Gamma-Poisson or Beta-Binomial distributions. DSS possess three characteristics to detect DMRs. First, spatial correlation. Proper utilization of the information from neighboring Cytosine sites can help improve estimation of methylation levels at each Cytosine site, and hence improve DMR detection. Second, the read depth of the Cytosine sites provides information on precision that can be exploited to improve statistical tests for DMR detection. Finally, the variance among biological replicates provides information necessary for a valid statistical test to detect DMRs，when there is no biological replicate, DSS combining data from nearby Cytosine sites and using them as ‘pseudo-replicates’ to estimate biological variance at specific locations.Sequence Alignment - The reference genome was firstly transformed into bisulfite-converted version (C-to-T and G-to-A converted) and then indexed using bowtie2 (Langmead B, 2012). Sequence reads were also transformed into fully bisulfite-converted versions (C-to-T and G-to-A converted) before they are aligned to similarly converted versions of the genome in a directional manner. Sequence reads that produce a unique best alignment from the two alignment processes (original top and bottom strand) are then compared to the normal genomic sequence and the methylation state of all cytosine positions in the read is inferredMetabolomicsUnknownTranscriptomicsGenomicsProteomicsIllumina MiSeqBisulfite-seqOryza sativa Japonica GroupLUIS ROMEROfalseHydrogen sulfide regulation of the DNA methylation profile of rice plants under drought stressTo investigate the effect of H2S on DNA methylation of plants submitted to drought stress, we performed transcriptome sequencing analysis in four experimental treatments: rice watered plants (control sample, C), rice plants watered with NaHS solution, as sulfide donor (sulfide treatment, S), rice plants subjected to drought stress (drought, D) and rice plants pretreated with NaHS and subjected to drought stress (S_D). Thus, 25-day-old plants grown in soil under physiological conditions were divided into two batches with one batch irrigated with water and the other with NaHS for 10 days, replacing the NaHS solution every two days. After this period, each batch was subsequently divided into two new batches and subjected to water irrigation or drought for another 15 additional days. At the end of the full treatment, four replicates of each condition were obtained that were processed independently2026-07-01T00:00:00Z2026-07-01T01:04:15.741Z2025-06-17T12:15:36.347ZE-MTAB-15231ERP173544E-MTAB-15224EFO_0002944EFO_0004170EFO_0003789EFO_0003753EFO_0004917EFO_0005518EFO_0003816EFO_0004184EFO_0003969