High-throughput single-cell RNA Sequencing is a powerful technique for gene expression profiling of complex and heterogeneous cellular populations such as the immune system. However, these methods only provide short-read sequences from one end of a cDNA template, making them poorly suited to the investigation of gene-regulatory events such as alternative RNA splicing, adaptive immune responses or somatic genome evolution. To address this challenge, we have developed a method that combines targeted long-read sequencing with short-read based transcriptome profiling of barcoded single cell libraries generated by droplet-based partitioning. We use Repertoire And Gene Expression by Sequencing (RAGE-Seq) on 7,138 lymphocytes sampled from the primary tumour and draining lymphnode of a breast cancer patient to accurately characterize full-length T-cell.
(TCR) and B-cell (BCR) receptor sequences and transcriptional profiles. With this method we show that somatic hypermutation, alternate splicing and clonal evolution of lymphocytes can be tracked across tissue compartments, defining signatures of expanded clones. Furthermore, we simultaneously demonstrate the utility of this methodology on human samples tagged with + 100 DNA barcoded antibody markers (CITE-seq), where we additionally show CD45 isoform splicing profile of individual immune cells.