title: GSM4775312: Rep 1 CD4 TN WT [RNA-Seq]; Mus musculus; RNA-Seq
library_construction_protocol: FACS sorted naive CD4+ T cells were sorted from lymph nodes and spleen. Naive CD4+ T cells were directly used for the RNA-seq protocol for the day 0 condition, while the remaining naive CD4+ T cells were activated in Th0 culture condition for 1 or 2 days. At day 1 and 2, live cells (DAPI-) were sorted and used for the RNA-seq protocol.  Total RNA extractions were performed using the RNeasy Plus Micro Kit (Qiagen 74034). Full length cDNA were generated from 5 ng of total RNA using Clontech SMART-Seq v4 Ultra Low Input RNA kit for Sequencing (Takara Bio Europe, Saint Germain en Laye, France) according to manufacturer's instructions with 9 cycles of PCR for cDNA amplification by Seq-Amp polymerase. Six hundreds pg of pre-amplified cDNA were then used as input for Tn5 transposon tagmentation by the Nextera XT DNA Library Preparation Kit (96 samples) (Illumina, San Diego, CA) followed by 12 cycles of library amplification. Following purification with Agencourt AMPure XP beads (Beckman-Coulter, Villepinte, France), the size and concentration of libraries were assessed by capillary electrophoreris.