Bisulfilte-Seq of cucumis sativus: temporal study
After samples DNA testing, positive control DNAs were added into the DNAs, and then they were fragmented into 200-400bp using Covaris S220. Next, terminal repairing, A-ligation, methylation sequencing adapters ligation were performed to the DNA fragments. And the final DNA library was ready after Bisulfite treatment (EZ DNA Methylation Gold Kit, Zymo Research. After BS treatment, unmethylated Cytosine will change into Uracil, while methylated Cytosine will stay unchanged), size selection and PCR amplification steps. The library kit 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. When read length is close to fragment size, the tail may also be observed toward the end of Read 1 data.