Cited by 136Single-cell and spatial transcriptomicsGenomics and Phylogenetic StudiesMolecular Biology Techniques and Applications

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Neuronal ambient RNA contamination causes misinterpreted and masked cell types in brain single-nuclei datasets

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Caglayan, Emre, Liu, Yuxiang, Konopka, Genevieve·2022·Neuron

DOI 10.1016/j.neuron.2022.09.010PMC PMC9789184

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Abstract

Ambient RNA contamination in single-cell and single-nuclei RNA sequencing (snRNA-seq) is a significant problem, but its consequences are poorly understood. Here, we show that ambient RNAs in brain snRNA-seq datasets have a nuclear or non-nuclear origin with distinct gene set signatures. Both ambient RNA signatures are predominantly neuronal, and we find that some previously annotated neuronal cell types are distinguished by ambient RNA contamination. We detect pervasive neuronal ambient RNA contamination in all glial cell types unless glia and neurons are physically separated prior to sequencing. We demonstrate that this contamination can be removed in silico and show that previous single-nuclei RNA-seq-based annotations of immature oligodendrocytes are glial nuclei contaminated with ambient RNAs. After ambient RNA removal, we detect rare, committed oligodendrocyte progenitor cells not annotated in most previous adult human brain datasets. Together, these results provide an in-depth analysis of ambient RNA contamination in brain single-nuclei datasets.

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Self-citation
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“Ambient RNA correction undoubtedly improves analysis outcomes in many settings [30,32,33,40].”
Influential
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eLife assessment: The gene expression landscape of the human locus coeruleus revealed by single-nucleus and spatially-resolved transcriptomics
Eisen, M. B. (2023) · eLife · doi:10.7554/elife.84628.1.sa4
Author Response: The gene expression landscape of the human locus coeruleus revealed by single-nucleus and spatially-resolved transcriptomics
Weber, L. M., Divecha, H. R., Tran, M. N., Kwon, S. H., Spangler, A., Montgomery, K. D., Tippani, M., Bharadwaj, R. A., Kleinman, J. E., Page, S. C., et al. (2023) · eLife · doi:10.7554/elife.84628.1.sa0
Reviewer #2 (Public Review): The gene expression landscape of the human locus coeruleus revealed by single-nucleus and spatially-resolved transcriptomics
(2023) · eLife · doi:10.7554/elife.84628.1.sa2
Reviewer #3 (Public Review): The gene expression landscape of the human locus coeruleus revealed by single-nucleus and spatially-resolved transcriptomics
(2023) · eLife · doi:10.7554/elife.84628.1.sa1

Authors: Caglayan, E., Liu, Y., & Konopka, G.
Affiliations: The University of Texas Southwestern Medical Center (US)
Corresponding author: Caglayan, Emre
ORCID: Caglayan, Emre
Liu, Yuxiang
Venue: Neuron
Publisher: Elsevier BV
Published: 2022-12
Volume: 110
Issue: 24
Pages: 4043-4056.e5
ISSN: 0896-6273 (ISSN-L)
DOI: 10.1016/j.neuron.2022.09.010
PMC: PMC9789184
OpenAlex: W4305016438
Cited by: 123
References: 60
Citations by year: 2026: 35 · 2025: 32 · 2024: 28 · 2023: 25 · 2022: 3
FWCI: 7.58
Type: journal article
Language: en
Open access: bronze
doi.org
Licence: www.elsevier.com/tdm/userlicense/1.0/ (tdm)
www.elsevier.com/legal/tdmrep-license (tdm)
www.elsevier.com/open-access/userlicense/1.0/ (vor, 385 d)
doi.org/10.15223/policy-017 (stm-asf)
doi.org/10.15223/policy-037 (stm-asf)
doi.org/10.15223/policy-012 (stm-asf)
doi.org/10.15223/policy-029 (stm-asf)
doi.org/10.15223/policy-004 (stm-asf)
Locations: Neuron (publishedVersion)
bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion, cc-by-nc-nd)
PubMed (publishedVersion)
PubMed Central (acceptedVersion)
Funders: James S. McDonnell Foundation
American Heart Association (915654)
University of Texas Southwestern Medical Center
National Institute of Mental Health (MH103517, MH126481, R01MH103517, R01MH126481)
National Human Genome Research Institute (R01HG011641, HG011641)
National Institute of Neurological Disorders and Stroke (NS115821, UF1NS115821)
Topics: Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Single-cell and spatial transcriptomics
Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Genomics and Phylogenetic Studies
Life Sciences › Biochemistry, Genetics and Molecular Biology › Molecular Biology › Molecular Biology Techniques and Applications
Keywords: single-nucleus RNA sequencing, brain cell types, oligodendrocyte precursor cells, cell type annotation, human brain
MeSH terms: Adult, Cell Nucleus, Humans, Neurons, RNA, RNA, Small Nuclear, Sequence Analysis, RNA, Solitary Nucleus, Single-Cell Analysis

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Neuronal ambient RNA contamination causes misinterpreted and masked cell types in brain single-nuclei datasets

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