GSM8942102: WGBS_LU_RAS_D0; Homo sapiens; Bisulfite-Seq
Total genomic DNA was extracted from rumen contents using the TIANGEN Magnetic Universal Genomic DNA Kit After samples DNA testing, positive control DNAs were added into the DNAs, and then they were fragmented into 200-400bp using Covaris S220（Covaris, USA）. Next, ssDNA fragements were Bisulfite treated. After BS treatment, unmethylated Cytosine will change into Uracil, while methylated Cytosine will stay unchanged. Then, methylation sequencing adapters ligation, size selection and PCR amplification steps were performed to the DNA fragments. The library kit (Accel-NGS Methyl-Seq DNA Library Kit(Swift, USA, Catalog #: 30096) adds a low complexity polynucleotide tail with an average length of 8 bases to the 3' end of each fragment during the addition of the first NGS adapter molecule. If these tails are not trimmed bioinformatically from the sequencing data, it is normal and expected to observe them at the beginning of Read 2 (R2). When read length is close to fragment size, the tail may also be observed toward the end of Read 1 (R1) data. Therebefore, it's highly recommended to trim 10 bases from the beginning of both R1 and R2 to eliminate any artifactual cytosine methylation introduced as a result of filling in overhangs during end repair steps of conventional dsDNA library preparation and low quality bases due to bisulfite treatment. Subsequently, library quality was assessed on the Agilent 5400 system（Agilent，USA） and quantified by QPCR(1.5 nM). Pair-end sequencing of sample was performed on Illumina platform (Illumina, USA).