title: GSM6250212: RNA Il17eGFP_d3_npTh17_all_2; Mus musculus; RNA-Seq
library_construction_protocol: 6,000 cells were sorted by FACS from the in vitro culture or draining lymph nodes and central nervous system of EAE mice into 5ul TCL buffer supplemented with 1% 2-mercaptoethanol and stored at -80°C until library preparation. For bulk RNA-seq, full-length RNA-seq libraries were prepared following a modified SMART-Seq2 protocol as previously described. The cDNA concentration was measured using the Quant-iT PicoGreen dsDNA Assay Kit (Thermo Fisher Scientific) and normalized to 0.25ng/ul. The cDNA libraries were prepared using the Nextera XT DNA Library Preparation kit (Illumina). Final libraries were confirmed to have a size of 500 bp using a Bioanalyzer (Agilent). Prior to sequencing, the uniquely barcoded libraries were pooled, normalized to 2 nM, and denatured using 0.1 N NaOH. Flow cell cluster amplification and sequencing were performed according to the manufacturer's protocols using the paired-end Illumina sequencing (38bp X 2) with 75 cycle Nextseq 500 high output V2 kit (Illumina).