Multiplexed droplet single-cell RNA-sequencing using natural genetic variation
Top Cited Papers
- 1 January 2018
- journal article
- research article
- Published by Springer Science and Business Media LLC in Nature Biotechnology
- Vol. 36 (1), 89-+
- https://doi.org/10.1038/nbt.4042
Abstract
Droplet single-cell RNA-sequencing (dscRNA-seq) has enabled rapid, massively parallel profiling of transcriptomes. However, assessing differential expression across multiple individuals has been hampered by inefficient sample processing and technical batch effects. Here we describe a computational tool, demuxlet, that harnesses natural genetic variation to determine the sample identity of each droplet containing a single cell (singlet) and detect droplets containing two cells (doublets). These capabilities enable multiplexed dscRNA-seq experiments in which cells from unrelated individuals are pooled and captured at higher throughput than in standard workflows. Using simulated data, we show that 50 single-nucleotide polymorphisms (SNPs) per cell are sufficient to assign 97% of singlets and identify 92% of doublets in pools of up to 64 individuals. Given genotyping data for each of eight pooled samples, demuxlet correctly recovers the sample identity of >99% of singlets and identifies doublets at rates consistent with previous estimates. We apply demuxlet to assess cell-type-specific changes in gene expression in 8 pooled lupus patient samples treated with interferon (IFN)-beta and perform eQTL analysis on 23 pooled samples.This publication has 53 references indexed in Scilit:
- Integrative genomic analysis of the human immune response to influenza vaccinationeLife, 2013
- Detecting and Estimating Contamination of Human DNA Samples in Sequencing and Array-Based Genotype DataAmerican Journal of Human Genetics, 2012
- STAR: ultrafast universal RNA-seq alignerBioinformatics, 2012
- Matrix eQTL: ultra fast eQTL analysis via large matrix operationsBioinformatics, 2012
- PrimerBank: a PCR primer database for quantitative gene expression analysis, 2012 updateNucleic Acids Research, 2011
- The variant call format and VCFtoolsBioinformatics, 2011
- Differential expression analysis for sequence count dataGenome Biology, 2010
- Balancing Selection Maintains a Form of ERAP2 that Undergoes Nonsense-Mediated Decay and Affects Antigen PresentationPLoS Genetics, 2010
- The Sequence Alignment/Map format and SAMtoolsBioinformatics, 2009
- PAnnBuilder: an R package for assembling proteomic annotation dataBioinformatics, 2009