GSM2551727: 90_healthy_Frontal Cortex; Homo sapiens; Bisulfite-Seq
DNA was isolated from 20–40 mg of sampled frontal cortex and cerebellum frozen tissue using the Wizard genomic DNA purification kit (Promega, Madison, WI, USA) as per manufacturers’ instructions, and re-suspended in TE buffer. Concentration was assessed using a Qubit 3.0 Fluorometer (ThermoFisher Scientific, Waltham, MA), and 50ng/ul of DNA in a 10ul volume was sent to the Mayo Clinic Medical Genome Facility in Rochester (Minnesota, USA) for processing and sequencing. RRBS library construction and sequencing was conducted at the Mayo Clinic Medical Genome Facility in Rochester (Minnesota, USA). First, the concentration and size distribution of the libraries were determined using Agilent (Santa Clara, CA, USA) Bioanalyzer DNA 1000 chip, Qubit fluorometry (Invitrogen, Carlsbad, CA), and qPCR. Libraries were pooled at equimolar concentrations and loaded onto a paired end flow cell at concentrations of 7-8 pM to generate cluster densities of 600,000-700,000/mm2 following Illumina’s standard protocol, using the Illumina cBot and HiSeq Paired end cluster kit version 3 (San Diego, CA, USA). Because bisulfite treated samples are unbalanced due to an under representation of C bases, a lane of PhiX genomic DNA was run to allow the Illumina data collection software to call bases correctly. Each lane of a HiSeq flow cell produces 21-39 Gbases of sequence. The flow cells were sequenced as 51 X 2 paired end reads on an Illumina HiSeq 2000 using TruSeq SBS sequencing kit version 3 and HiSeq data collection version 2.0.12.0 software. Base-calling was performed using Illumina’s RTA version 1.17.21.3.
GEO Accession
GSM2551727