title: GSM3444176: SP-4-ATCCATG_S4_L001_R1_001: Isotype_1 (Iso); Mus musculus; RNA-Seq
library_construction_protocol: Live CD45+CD3+CD8+Spas-1+Spas-2- T cells were sorted from draining lymph nodes from TRAMP-C2 bearing mice on day 28 after treatment. Cells from the five mice from each treatment arm were pooled and split into triplicates. RNA was extracted from sorted Live CD45+CD3+CD8+Spas-1+Spas-2- T cells using an Ambion micro RNA isolation kit (AM1931) according to the manufacturer's protocol. Bulk RNA sequencing was carried out on 0.25ng of RNA in triplicates from each treatment type using the protocol as previously described (Picelli et al., 2014) with the following modifications. The TSO, Oligo-dT30VN, and ISPCR primers were biotinylated on the 5' end. 21 cycles of pre-amplification were found to be optimal. Nextera adapters were added and the libraries were pooled and sequenced on an Illumina Hiseq 4000. Approximately 48,710 reads were obtained per library. The Fastq sequences were aligned and quantitated using the CLC genomics workbench RNA-Seq analysis tools (Qiagen Bioinformatics).