title: Droplet barcoding for single-cell transcriptomics applied to embryonic stem cells.
abstract: It has long been the dream of biologists to map gene expression at the single-cell level. With such data one might track heterogeneous cell sub-populations, and infer regulatory relationships between genes and pathways. Recently, RNA sequencing has achieved single-cell resolution. What is limiting is an effective way to routinely isolate and process large numbers of individual cells for quantitative in-depth sequencing. We have developed a high-throughput droplet-microfluidic approach for barcoding the RNA from thousands of individual cells for subsequent analysis by next-generation sequencing. The method shows a surprisingly low noise profile and is readily adaptable to other sequencing-based assays. We analyzed mouse embryonic stem cells, revealing in detail the population structure and the heterogeneous onset of differentiation after leukemia inhibitory factor (LIF) withdrawal. The reproducibility of these high-throughput single-cell data allowed us to deconstruct cell populations and infer gene expression relationships. VIDEO ABSTRACT.
authors.affiliation: Department of Systems Biology, Harvard Medical School, Boston, MA 02115, USA.
authors.forename: Allon M
authors.initials: AM
authors.lastname: Klein
authors.affiliation: School of Engineering and Applied Sciences (SEAS), Harvard University, Cambridge, MA 02138, USA; Vilnius University Institute of Biotechnology, Vilnius LT-02241, Lithuania.
authors.forename: Linas
authors.initials: L
authors.lastname: Mazutis
authors.affiliation: School of Engineering and Applied Sciences (SEAS), Harvard University, Cambridge, MA 02138, USA.
authors.forename: Ilke
authors.initials: I
authors.lastname: Akartuna
authors.affiliation: Department of Systems Biology, Harvard Medical School, Boston, MA 02115, USA.
authors.forename: Naren
authors.initials: N
authors.lastname: Tallapragada
authors.affiliation: Department of Systems Biology, Harvard Medical School, Boston, MA 02115, USA; Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA 02138, USA; Harvard Stem Cell Institute, Harvard University, Cambridge, MA 02138, USA.
authors.forename: Adrian
authors.initials: A
authors.lastname: Veres
authors.affiliation: Department of Systems Biology, Harvard Medical School, Boston, MA 02115, USA.
authors.forename: Victor
authors.initials: V
authors.lastname: Li
authors.affiliation: Department of Systems Biology, Harvard Medical School, Boston, MA 02115, USA.
authors.forename: Leonid
authors.initials: L
authors.lastname: Peshkin
authors.affiliation: School of Engineering and Applied Sciences (SEAS), Harvard University, Cambridge, MA 02138, USA. Electronic address: weitz@seas.harvard.edu.
authors.forename: David A
authors.initials: DA
authors.lastname: Weitz
authors.affiliation: Department of Systems Biology, Harvard Medical School, Boston, MA 02115, USA. Electronic address: marc@hms.harvard.edu.
authors.forename: Marc W
authors.initials: MW
authors.lastname: Kirschner
mesh_terms: D000818:Animals; D053595:Embryonic Stem Cells / Q000166:cytology* / Q000378:metabolism; D020869:Gene Expression Profiling / Q000379:methods*; D059014:High-Throughput Nucleotide Sequencing; D051379:Mice; D046210:Microfluidic Analytical Techniques; D017423:Sequence Analysis, RNA / Q000379:methods; D059010:Single-Cell Analysis / Q000379:methods*