title: GSM3731227: R3; Oryza sativa; Bisulfite-Seq
library_construction_protocol: DNA was extracted using the DNeasy plant mini kit  (Qiagen) according to manufacturer's instructions and dissolved in DNase free water. DNA concentration was quantified using Quant-it PicoGreen dsDNA Assay kit (Thermo Fisher  Scientific) and 100 ng of DNA per sample was sheared to 400 bp fragments using Covaris S2  Focused-ultrasonicator Sonicator settings were as following: duty cycles of 10 %, intensity of  4, cycles/burst of 200 for 65 seconds. Fragmented DNA was cleaned and concentrated using the DNA clean and concentrator kit (Zymo Research). Bisulfite conversion was performed by using the EZ DNA methylation Gold kit (Zymo Research). Evaluation of bisulfite conversion efficiency was done by shearing the unmethylated Lambda DNA (48502 bp) as above. Size distribution was verified using the high sensitivity DNA chip of Bioanalyzer 2100 (Agilent technologies). The preparation of sequencing libraries from bisulfite-converted DNA was performed using Accel-NGS Methyl-Seq DNA library kit according to manufacturer's instructions (Swift Biosciences). The quality of DNA was again checked by using the high sensitivity DNA chip of Bioanalyzer 2100 (Agilent technologies). After gel purification, qPCR was performed to determine the concentration of each sample. Finally, a 2.1 pM library was loaded on a single read flow cell, and sequenced using an Illumina Nextseq500 (single end, 76 cycles).
attributes.tag: GEO Accession
attributes.value: GSM3731227