title: GSM7328883: Rat CD4+ T-MBP cells with GW library, parenchyma, rep R2; Rattus norvegicus; OTHER
library_construction_protocol: The spinal cord parenchyma was dissected and homogenized by passing through a metal strainer. Lymphocytes were isolated using a 30%/64% Percoll gradient and centrifugation at 1200g, room temperature for 30min with mild acceleration and brake. Lymphocytes were collected from the interface. Genomic DNA (gDNA) from lymphocytes or sorted TMBP cells was isolated with the DNeasy Blood and Tissue Kit (Qiagen). A one-step PCR amplification was performed with Q5 High Fidelity DNA Polymerase by using 2.5µg of gDNA per reaction with Fwd-Lib (mix of 8 staggered primers) and Rev-Lib (consists of 8bp of unique barcode) primers for a total of 24 cycles. Illumina adapters were introduced together with the amplification primers. All primer sequences are listed in Supplementary Table 1. The amplified DNA amplicons were purified with SPRIselect (Beckman Coulter) with a ratio of 1:0.8 (DNA to beads) and eluted in nuclease-free water. The presence of ~250bp DNA amplicons was confirmed and the concentration was measured with Agilent Bioanalyzer on DNA 1000 Chips (5067-1504).