Single cell and bulk RNAseq profiles of thymic pro-T cells

The single cell and bulk expression profiles of early pro T-cell progenitors from mouse thymus Examination of expression profiles of thymic ETP-DN2 population with bulk RNAseq and single cell RNAseq. 10X: Droplet-based 3′ end massively parallel single-cell RNA sequencing droplets were prepared using Chromium Single Cell 3′ Reagent Kits v2 according to manufacturer’s protocol (10x Genomics). C1: Microfluidic-based full length mRNA scRNAseq were performed with Single-Cell mRNA Seq IFC 5–10 µm chip, and Smartseq 2 chemistry. Bulk: Bulk RNAseq from FACS sorted early T cell populations were performed. RNAeasy MicroKit (Qiagen) were used to purify the total RNA according to manufacturer’s recommendations. Purified RNA samples have been processed with and without SMART-Seq v4 Ultra Low Input RNA Kit(Takara #634888) for pre-amplification, and Nextera® XT library preparation kits for Illumina sequencing. Samples without pre-amp were constructed using NEBNext Ultra RNA Library Prep Kit for Illumina (NEB #E7530) from ~1 μg of total RNA following manufacturer’s instructions. Libraries were sequenced on Illumina HiSeq2500 in single read mode with the read length of 50-51 nt following manufacturer's instructions. Base calls were performed with RTA 1.13.48.0 followed by conversion to FASTQ with bcl2fastq 1.8.4 and produced approximately 30 million reads per sample.