Bisulfite_Seq_I50M1_3_Internodes_ 50 zeb_ 1 month soil_ rep3
Genome-wide DNA Methylation (Bisulphite-seq) (Novogene): DNA quantification and qualification: Genomic DNA degradation and contamination was monitored on agarose gels. DNA concentration was measured using Qubit DNA Assay Kit in Qubit 2.0 Flurometer (Life Technologies, CA, USA). Library preparation and quantification: A total amount of 5.2 microgram genomic DNA spiked with 26 ng lambda DNA were fragmented by sonication to 200-400 bp with Covaris S220, followed by end repair and adenylation. Cytosine-methylated barcodes were ligated to sonicated DNA as per manufacturers instructions. Then these DNA fragments were treated twice with bisulfite using EZ DNA Methylation-GoldTM Kit (Zymo Research). And the resulting single-strand DNA fragments were PCR amplificated using KAPA HiFi HotStart Uracil + ReadyMix (2X). Library concentration was quantified by Qubit 2.0 Flurometer (Life Technologies, CA, USA) and quantitative PCR, and the insert size was checkedBoinoaAngaliyleznetr 2100 system. Clustering and sequencing: The clustering of the index-coded samples was performed on a cBot Cluster Generation System using PE Cluster Kit cBot-HS (Illumina) according to the manufacturers instructions. After cluster generation, the library preparations were sequenced on an Illumina platform and 150 bp paired-end reads were generated. Image analysis and base calling were performed with the standard Illumina pipeline, and finally 150  bp paired-end reads were generated.