LOTR-Seq: A protocol for large-scale simultaneous single-cell long-read genotyping of transcripts.

Single-cell RNA sequencing enables analysis of intra-tumoral transcriptional heterogeneity but has limited coverage of mutational hotspots. Here, we present LOTR-Seq, a protocol combining short-read whole-transcriptome analysis with long-read genotyping. We describe steps for hematopoietic stem cell isolation from human bone marrow samples, generation of full-length single-cell barcoded cDNA, target enrichment, and long-read sequencing for variant calling. For complete details on the use and execution of this protocol, please refer to Grabek et al.1.