GSM6691935: setB_control2_control1; Mus musculus; RNA-Seq
C4+ T cells were isolated from mouse spleens using MACS beads for positive selection. Cells were activated using anti-CD3 and anti-CD28 for 40 hrs, then transduced with combined concentrated ecotropic retrovirus supernatants for circIkzf1 (30), control1(28), Ikzf1 (38) and control2 (8) using spin infection. Cells were then cultured until day 5, when they were FACS sorted for the expression of the two transduction markers hCD4 and GFP. Sorted cells were subjected to single cell capturing according to the 10x Genomics 5' GEX protocol. The purified cDNA was amplified using 14 cycles of PCR and then used for 5' gene expression (GEX). Through fragmentation, adapter ligation and index PCR the final libraries were obtained. The quality of single cell 5' GEX was assessed by Qubit quantification and fragment analysis (HS DNA Kit, Agilent).The sequencing was performed on a NextSeq500 device (Illumina) with the recommended sequencing conditions for 5' GEX libraries (read1: 26nt, read2: 98nt, index1: 8nt, index2:0).