title: Low-coverage single-cell mRNA sequencing reveals cellular heterogeneity and activated signaling pathways in developing cerebral cortex.
abstract: Large-scale surveys of single-cell gene expression have the potential to reveal rare cell populations and lineage relationships but require efficient methods for cell capture and mRNA sequencing. Although cellular barcoding strategies allow parallel sequencing of single cells at ultra-low depths, the limitations of shallow sequencing have not been investigated directly. By capturing 301 single cells from 11 populations using microfluidics and analyzing single-cell transcriptomes across downsampled sequencing depths, we demonstrate that shallow single-cell mRNA sequencing (~50,000 reads per cell) is sufficient for unbiased cell-type classification and biomarker identification. In the developing cortex, we identify diverse cell types, including multiple progenitor and neuronal subtypes, and we identify EGR1 and FOS as previously unreported candidate targets of Notch signaling in human but not mouse radial glia. Our strategy establishes an efficient method for unbiased analysis and comparison of cell populations from heterogeneous tissue by microfluidic single-cell capture and low-coverage sequencing of many cells.
authors.affiliation: 1] Eli and Edythe Broad Center of Regeneration Medicine and Stem Cell Research, University of California, San Francisco, San Francisco, California, USA. [2] Department of Neurology, University of California, San Francisco, San Francisco, California, USA. [3].
authors.forename: Alex A
authors.initials: AA
authors.lastname: Pollen
authors.affiliation: 1] Eli and Edythe Broad Center of Regeneration Medicine and Stem Cell Research, University of California, San Francisco, San Francisco, California, USA. [2] Department of Neurology, University of California, San Francisco, San Francisco, California, USA. [3].
authors.forename: Tomasz J
authors.initials: TJ
authors.lastname: Nowakowski
authors.affiliation: 1] Fluidigm Corporation, South San Francisco, California, USA. [2].
authors.forename: Joe
authors.initials: J
authors.lastname: Shuga
authors.affiliation: 1] Fluidigm Corporation, South San Francisco, California, USA. [2].
authors.forename: Xiaohui
authors.initials: X
authors.lastname: Wang
authors.affiliation: Fluidigm Corporation, South San Francisco, California, USA.
authors.forename: Anne A
authors.initials: AA
authors.lastname: Leyrat
authors.affiliation: 1] Eli and Edythe Broad Center of Regeneration Medicine and Stem Cell Research, University of California, San Francisco, San Francisco, California, USA. [2] Department of Neurology, University of California, San Francisco, San Francisco, California, USA.
authors.forename: Jan H
authors.initials: JH
authors.lastname: Lui
authors.affiliation: Fluidigm Corporation, South San Francisco, California, USA.
authors.forename: Nianzhen
authors.initials: N
authors.lastname: Li
authors.affiliation: Fluidigm Corporation, South San Francisco, California, USA.
authors.forename: Lukasz
authors.initials: L
authors.lastname: Szpankowski
authors.affiliation: Fluidigm Corporation, South San Francisco, California, USA.
authors.forename: Brian
authors.initials: B
authors.lastname: Fowler
authors.affiliation: Fluidigm Corporation, South San Francisco, California, USA.
authors.forename: Peilin
authors.initials: P
authors.lastname: Chen
authors.affiliation: Fluidigm Corporation, South San Francisco, California, USA.
authors.forename: Naveen
authors.initials: N
authors.lastname: Ramalingam
authors.affiliation: Fluidigm Corporation, South San Francisco, California, USA.
authors.forename: Gang
authors.initials: G
authors.lastname: Sun
authors.affiliation: Fluidigm Corporation, South San Francisco, California, USA.
authors.forename: Myo
authors.initials: M
authors.lastname: Thu
authors.affiliation: Fluidigm Corporation, South San Francisco, California, USA.
authors.forename: Michael
authors.initials: M
authors.lastname: Norris
authors.affiliation: Fluidigm Corporation, South San Francisco, California, USA.
authors.forename: Ronald
authors.initials: R
authors.lastname: Lebofsky
authors.affiliation: Fluidigm Corporation, South San Francisco, California, USA.
authors.forename: Dominique
authors.initials: D
authors.lastname: Toppani
authors.affiliation: Fluidigm Corporation, South San Francisco, California, USA.
authors.forename: Darnell W
authors.initials: DW
authors.lastname: Kemp
authors.affiliation: Fluidigm Corporation, South San Francisco, California, USA.
authors.forename: Michael
authors.initials: M
authors.lastname: Wong
authors.affiliation: Fluidigm Corporation, South San Francisco, California, USA.
authors.forename: Barry
authors.initials: B
authors.lastname: Clerkson
authors.affiliation: Fluidigm Corporation, South San Francisco, California, USA.
authors.forename: Brittnee N
authors.initials: BN
authors.lastname: Jones
authors.affiliation: Fluidigm Corporation, South San Francisco, California, USA.
authors.forename: Shiquan
authors.initials: S
authors.lastname: Wu
authors.affiliation: Fluidigm Corporation, South San Francisco, California, USA.
authors.forename: Lawrence
authors.initials: L
authors.lastname: Knutsson
authors.affiliation: Fluidigm Corporation, South San Francisco, California, USA.
authors.forename: Beatriz
authors.initials: B
authors.lastname: Alvarado
authors.affiliation: Fluidigm Corporation, South San Francisco, California, USA.
authors.forename: Jing
authors.initials: J
authors.lastname: Wang
authors.affiliation: Fluidigm Corporation, South San Francisco, California, USA.
authors.forename: Lesley S
authors.initials: LS
authors.lastname: Weaver
authors.affiliation: Fluidigm Corporation, South San Francisco, California, USA.
authors.forename: Andrew P
authors.initials: AP
authors.lastname: May
authors.affiliation: Fluidigm Corporation, South San Francisco, California, USA.
authors.forename: Robert C
authors.initials: RC
authors.lastname: Jones
authors.affiliation: Fluidigm Corporation, South San Francisco, California, USA.
authors.forename: Marc A
authors.initials: MA
authors.lastname: Unger
authors.affiliation: 1] Eli and Edythe Broad Center of Regeneration Medicine and Stem Cell Research, University of California, San Francisco, San Francisco, California, USA. [2] Department of Neurology, University of California, San Francisco, San Francisco, California, USA.
authors.forename: Arnold R
authors.initials: AR
authors.lastname: Kriegstein
authors.affiliation: Fluidigm Corporation, South San Francisco, California, USA.
authors.forename: Jay A A
authors.initials: JA
authors.lastname: West
mesh_terms: D000818:Animals; D002540:Cerebral Cortex / Q000254:growth & development* / Q000378:metabolism; D019295:Computational Biology / Q000379:methods*; D004867:Equipment Design; D020869:Gene Expression Profiling / Q000379:methods*; D006801:Humans; D051379:Mice; D046210:Microfluidic Analytical Techniques; D012333:RNA, Messenger / Q000032:analysis* / Q000235:genetics / Q000378:metabolism; D017423:Sequence Analysis, RNA / Q000379:methods*; D015398:Signal Transduction / Q000235:genetics* / Q000502:physiology
chemical_list: D012333:RNA, Messenger
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