High-Throughput Single-Cell Sequencing with Linear Amplification

Conventional methods for single-cell genome sequencing are limited with respect to uniformity and throughput. Here, we describe sci-L3, a single-cell sequencing method that combines combinatorial indexing (sci-) and linear (L) amplification. The sci-L3 method adopts a 3-level (3) indexing scheme that minimizes amplification biases while enabling exponential gains in throughput. We demonstrate the generalizability of sci-L3 with proof-of-concept demonstrations of single-cell whole-genome sequencing (sci-L3-WGS), targeted sequencing (sci-L3-target-seq), and a co-assay of the genome and transcriptome (sci-L3-RNA/DNA). We apply sci-L3-WGS to profile the genomes of >10,000 sperm and sperm precursors from F1 hybrid mice, mapping 86,786 crossovers and characterizing rare chromosome mis-segregation events in meiosis, including instances of whole-genome equational chromosome segregation. We anticipate that sci-L3 assays can be applied to fully characterize recombination landscapes, to couple CRISPR perturbations and measurements of genome stability, and to other goals requiring high-throughput, high-coverage single-cell sequencing. [Display omitted] •Flexible method for single-cell whole genomes, targeted sequencing, and RNA/DNA co-assay•Potential throughput of 1 million cells by 3-level single-cell combinatorial indexing•Linear amplification by in vitro transcription for uniform coverage of the genome•Discovery of whole-genome equational chromosome segregation in mouse meiosis I Yin et al. developed sci-L3, which combines a scalable single-cell barcoding scheme with linear amplification. The method flexibly enables single-cell whole-genome sequencing, targeted DNA sequencing, or concurrent profiling of the genome and transcriptome. With sci-L3, the authors discovered mitotic-like whole-genome chromosome segregation in male mouse meiosis I.