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Status |
Public on Oct 19, 2022 |
Title |
Ambient RNA analysis reveals misinterpreted and masked cell types in brain single-nuclei datasets |
Platform organism |
Homo sapiens |
Sample organisms |
Homo sapiens; Mus musculus |
Experiment type |
Expression profiling by high throughput sequencing Third-party reanalysis
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Summary |
Ambient RNA contamination in single-cell RNA sequencing (RNA-seq) is a significant problem, but its consequences are poorly understood. Here, we show that ambient RNAs in brain single-nuclei RNA-seq can have a nuclear or extra-nuclear origin, and each origin results in a distinct gene set signature. Both ambient RNA signatures are predominantly neuronal and we find that some previously annotated neuronal cell types are distinguished by ambient RNA. Strikingly, we also 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 using existing tools. We also show that previous annotations of immature oligodendrocytes are likely glial cells contaminated with ambient RNAs. After ambient RNA removal, we can detect extremely rare committed oligodendrocyte progenitor cells, which were infrequently annotated in previously published adult human brain datasets. Together, these results provide an in-depth analysis of ambient RNA contamination in brain single-cell datasets.
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Overall design |
Data were reanalzyzed from raw fastq files with or without CellBender. Cells were annotated based on their cell type and glial cells were additionally annotated for ambient RNA contamination based on ambient RNA marker gene enrichment. Oligodendrocyte progenitor cells were additionally clustered and annotated. --------------------------------------- Reanalyzed data includes PRJNA434002 (SRR9262927-SRR9262957), GSE192773, GSE97930, and GSE198640)
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Contributor(s) |
Caglayan E, Liu Y, Konopka G |
Citation(s) |
36240767 |
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Submission date |
Mar 18, 2022 |
Last update date |
Oct 19, 2022 |
Contact name |
Genevieve Konopka |
E-mail(s) |
genevieve.konopka@utsouthwestern.edu
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Organization name |
University of Texas Southwestern Medical Center
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Department |
Neuroscience
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Lab |
Konopka
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Street address |
5323 Harry Hines Blvd.
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City |
Dallas |
State/province |
Texas |
ZIP/Postal code |
75390 |
Country |
USA |
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Platforms (2) |
GPL16791 |
Illumina HiSeq 2500 (Homo sapiens) |
GPL24676 |
Illumina NovaSeq 6000 (Homo sapiens) |
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Samples (4)
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Relations |
Reanalysis of |
GSM5765186 |
Reanalysis of |
GSM5765187 |
Reanalysis of |
GSM5765188 |
Reanalysis of |
GSM5765189 |
Reanalysis of |
GSM2581267 |
Reanalysis of |
GSM2581268 |
Reanalysis of |
GSM2581269 |
Reanalysis of |
GSM2581270 |
Reanalysis of |
GSM2581271 |
Reanalysis of |
GSM2581272 |
Reanalysis of |
GSM2581273 |
Reanalysis of |
GSM2581274 |
Reanalysis of |
GSM2581275 |
Reanalysis of |
GSM2581276 |
Reanalysis of |
GSM2581277 |
Reanalysis of |
GSM2581278 |
Reanalysis of |
GSM2581279 |
Reanalysis of |
GSM2581280 |
Reanalysis of |
GSM2581281 |
Reanalysis of |
GSM2581282 |
Reanalysis of |
GSM2581283 |
Reanalysis of |
GSM2581284 |
Reanalysis of |
GSM2581285 |
Reanalysis of |
GSM2581286 |
Reanalysis of |
GSM2581287 |
Reanalysis of |
GSM2581288 |
Reanalysis of |
GSM2581289 |
Reanalysis of |
GSM2581290 |
Reanalysis of |
GSM2581291 |
Reanalysis of |
GSM2581292 |
Reanalysis of |
GSM2581293 |
Reanalysis of |
GSM2581294 |
Reanalysis of |
GSM2581295 |
Reanalysis of |
GSM2581296 |
Reanalysis of |
GSM2581297 |
Reanalysis of |
GSM2581298 |
Reanalysis of |
GSM2734257 |
Reanalysis of |
GSM2734258 |
Reanalysis of |
GSM2734259 |
Reanalysis of |
GSM2734263 |
Reanalysis of |
GSM2734264 |
Reanalysis of |
GSM5954173 |
Reanalysis of |
GSM5954174 |
Reanalysis of |
GSM5954175 |
Reanalysis of |
GSM5954176 |
BioProject |
PRJNA865472 |
Supplementary file |
Size |
Download |
File type/resource |
GSE198951_NSD1_CellBender_Glia.RDS.gz |
141.6 Mb |
(ftp)(http) |
RDS |
GSE198951_NSD1_NoCellBender_Glia.RDS.gz |
159.5 Mb |
(ftp)(http) |
RDS |
GSE198951_NSD1_OPC_Seurat.RDS.gz |
1.8 Gb |
(ftp)(http) |
RDS |
GSE198951_NSD1_intronicRatios.txt.gz |
15.5 Mb |
(ftp)(http) |
TXT |
GSE198951_NSD2_CellBender_Glia.RDS.gz |
41.7 Mb |
(ftp)(http) |
RDS |
GSE198951_NSD2_NoCellBender_Glia.RDS.gz |
560.5 Mb |
(ftp)(http) |
RDS |
GSE198951_NSD2_OPC_Seurat.RDS.gz |
89.3 Mb |
(ftp)(http) |
RDS |
GSE198951_NSD2_intronicRatios.txt.gz |
1.4 Mb |
(ftp)(http) |
TXT |
GSE198951_README_GEO.txt |
757 b |
(ftp)(http) |
TXT |
GSE198951_SD1_CellBender_Glia.RDS.gz |
13.9 Mb |
(ftp)(http) |
RDS |
GSE198951_SD1_NoCellBender_Glia.RDS.gz |
16.7 Mb |
(ftp)(http) |
RDS |
GSE198951_SD1_OPC_Seurat.RDS.gz |
24.7 Mb |
(ftp)(http) |
RDS |
GSE198951_SD1_intronicRatios.txt.gz |
1.1 Mb |
(ftp)(http) |
TXT |
GSE198951_SD2_CellBender_Glia.RDS.gz |
114.5 Mb |
(ftp)(http) |
RDS |
GSE198951_SD2_NoCellBender_Glia.RDS.gz |
26.0 Mb |
(ftp)(http) |
RDS |
SRA Run Selector |
Raw data not applicable for this record |
Processed data provided as supplementary file |