High-content Single-cell Combinatorial Indexing

Overview

Journal Nat Biotechnol

Specialty Biotechnology

Date 2021 Jul 6

PMID 34226710

Citations 36

Authors

Ryan M Mulqueen

Dmitry Pokholok

Brendan L OConnell

Casey A Thornton

Fan Zhang

Brian J ORoak

Jason Link

Galip Gurkan Yardimci

Rosalie C Sears

Frank J Steemers

Andrew C Adey

Affiliations

Soon will be listed here.

Abstract

Single-cell combinatorial indexing (sci) with transposase-based library construction increases the throughput of single-cell genomics assays but produces sparse coverage in terms of usable reads per cell. We develop symmetrical strand sci ('s3'), a uracil-based adapter switching approach that improves the rate of conversion of source DNA into viable sequencing library fragments following tagmentation. We apply this chemistry to assay chromatin accessibility (s3-assay for transposase-accessible chromatin, s3-ATAC) in human cortical and mouse whole-brain tissues, with mouse datasets demonstrating a six- to 13-fold improvement in usable reads per cell compared with other available methods. Application of s3 to single-cell whole-genome sequencing (s3-WGS) and to whole-genome plus chromatin conformation (s3-GCC) yields 148- and 14.8-fold improvements, respectively, in usable reads per cell compared with sci-DNA-sequencing and sci-HiC. We show that s3-WGS and s3-GCC resolve subclonal genomic alterations in patient-derived pancreatic cancer cell lines. We expect that the s3 platform will be compatible with other transposase-based techniques, including sci-MET or CUT&Tag.

Citing Articles

Generalization of the sci-L3 method to achieve high-throughput linear amplification for replication template strand sequencing, genome conformation capture, and the joint profiling of RNA and chromatin accessibility.

Chovanec P, Yin Y Nucleic Acids Res. 2025; 53(4).

PMID: 39997216 PMC: 11851118. DOI: 10.1093/nar/gkaf101.

Single-Cell Hi-C Technologies and Computational Data Analysis.

Dautle M, Chen Y Adv Sci (Weinh). 2025; 12(9):e2412232.

PMID: 39887949 PMC: 11884588. DOI: 10.1002/advs.202412232.

Exploring a pico-well based scRNA-seq method (HIVE) for simplified processing of equine bronchoalveolar lavage cells.

Fegraeus K, Riihimaki M, Nordlund J, Akula S, Wernersson S, Raine A PLoS One. 2025; 20(1):e0317343.

PMID: 39854349 PMC: 11760581. DOI: 10.1371/journal.pone.0317343.

Droplet-based high-throughput 3D genome structure mapping of single cells with simultaneous transcriptomics.

Wu H, Wang M, Zheng Y, Xie X Cell Discov. 2025; 11(1):8.

PMID: 39837831 PMC: 11751028. DOI: 10.1038/s41421-025-00770-8.

Advances and applications in single-cell and spatial genomics.

Wang J, Ye F, Chai H, Jiang Y, Wang T, Ran X Sci China Life Sci. 2025; .

PMID: 39792333 DOI: 10.1007/s11427-024-2770-x.

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