GSM3581218: Control 2_hMeDIP-seq; Bubalus bubalis; MeDIP-Seq
The genomic DNA of DCs were used for hMeDIP and library construction; it was isolated from each individual sample using a DNeasy Blood & Tissue Kit (Qiagen, USA). Quantification and quality assurance were evaluated using the NanoDrop 1000 (Agilent, USA). To generate the library, the products of the hMeDIP, as well as the supernatant input DNA, were purified using Qiagen MinElute columns and eluted in 16 μL of elution buffer (Qiagen, USA). Fourteen cycles of PCR were performed on 5 μL of the immunoprecipitated DNA using the single-end Illumina PCR primers. The resulting reactions were purified with Qiagen MinElute columns; then, a final size selection (~300-600 bp) was performed using AMPure XP beads. Quality control of the libraries was performed by the Agilent 2100 Bioanalyzer using the Agilent DNA 1000 chip kitChip Kit (Agilent, USA). An aliquot of each library was diluted in EB to 5 ng/μL, and 1 μL was used in real-time PCR reactions to confirm enrichment of the hydroxymethylated region.