title: Deep sequencing reveals cell-type-specific patterns of single-cell transcriptome variation.
abstract: BACKGROUND
  Differentiation of metazoan cells requires execution of different gene expression programs but recent single-cell transcriptome profiling has revealed considerable variation within cells of seeming identical phenotype. This brings into question the relationship between transcriptome states and cell phenotypes. Additionally, single-cell transcriptomics presents unique analysis challenges that need to be addressed to answer this question.
  
  
  RESULTS
  We present high quality deep read-depth single-cell RNA sequencing for 91 cells from five mouse tissues and 18 cells from two rat tissues, along with 30 control samples of bulk RNA diluted to single-cell levels. We find that transcriptomes differ globally across tissues with regard to the number of genes expressed, the average expression patterns, and within-cell-type variation patterns. We develop methods to filter genes for reliable quantification and to calibrate biological variation. All cell types include genes with high variability in expression, in a tissue-specific manner. We also find evidence that single-cell variability of neuronal genes in mice is correlated with that in rats consistent with the hypothesis that levels of variation may be conserved.
  
  
  CONCLUSIONS
  Single-cell RNA-sequencing data provide a unique view of transcriptome function; however, careful analysis is required in order to use single-cell RNA-sequencing measurements for this purpose. Technical variation must be considered in single-cell RNA-sequencing studies of expression variation. For a subset of genes, biological variability within each cell type appears to be regulated in order to perform dynamic functions, rather than solely molecular noise.
authors.affiliation: Department of Genomics and Computational Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA. hdueck@mail.upenn.edu.
authors.forename: Hannah
authors.initials: H
authors.lastname: Dueck
authors.affiliation: Department of Biology, School of Arts and Sciences, University of Pennsylvania, 301A/B Lynch Laboratory, 433 S University Avenue, Philadelphia, PA, 19104, USA. mugdhak@pcbi.upenn.edu.
authors.forename: Mugdha
authors.initials: M
authors.lastname: Khaladkar
authors.affiliation: Department of Pharmacology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA. sean.taekyung@gmail.com.|Current address: Allen Institute for Brain Science, Seattle, WA, USA. sean.taekyung@gmail.com.
authors.forename: Tae Kyung
authors.initials: TK
authors.lastname: Kim
authors.affiliation: Department of Pharmacology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA. jspaethl@mail.med.upenn.edu.
authors.forename: Jennifer M
authors.initials: JM
authors.lastname: Spaethling
authors.affiliation: Department of Biology, School of Arts and Sciences, University of Pennsylvania, 301A/B Lynch Laboratory, 433 S University Avenue, Philadelphia, PA, 19104, USA. chantalfedde@gmail.com.
authors.forename: Chantal
authors.initials: C
authors.lastname: Francis
authors.affiliation: Department of Pediatrics, Harvard Medical School, Boston, MA, USA. sangita.suresh@gmail.com.|Department of Cardiology, Boston Children's Hospital, Boston, MA, USA. sangita.suresh@gmail.com.
authors.forename: Sangita
authors.initials: S
authors.lastname: Suresh
authors.affiliation: Department of Biology, School of Arts and Sciences, University of Pennsylvania, 301A/B Lynch Laboratory, 433 S University Avenue, Philadelphia, PA, 19104, USA. safisher@sas.upenn.edu.
authors.forename: Stephen A
authors.initials: SA
authors.lastname: Fisher
authors.affiliation: Department of Cell and Developmental Biology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA. sealep@mail.med.upenn.edu.
authors.forename: Patrick
authors.initials: P
authors.lastname: Seale
authors.affiliation: Department of Anesthesiology, The Children's Hospital of Philadelphia Research Institute, Philadelphia, PA, USA. BECKS@email.chop.edu.
authors.forename: Sheryl G
authors.initials: SG
authors.lastname: Beck
authors.affiliation: The Department of Chemical Physiology, The Scripps Research Institute, La Jolla, CA, USA. tbartfai@scripps.edu.
authors.forename: Tamas
authors.initials: T
authors.lastname: Bartfai
authors.affiliation: Department of Pediatrics, Harvard Medical School, Boston, MA, USA. Bernhard.Kuhn2@CHP.edu.|Department of Cardiology, Boston Children's Hospital, Boston, MA, USA. Bernhard.Kuhn2@CHP.edu.|Harvard Stem Cell Institute, Cambridge, MA, USA. Bernhard.Kuhn2@CHP.edu.|Department of Stem Cell and Regenerative Biology, Harvard University, Cambridge, MA, USA. Bernhard.Kuhn2@CHP.edu.|Current address: Richard King Mellon Institute for Pediatric Research, Children's Hospital of Pittsburgh of UPMC, Pittsburgh, PA, USA. Bernhard.Kuhn2@CHP.edu.
authors.forename: Bernhard
authors.initials: B
authors.lastname: Kuhn
authors.affiliation: Department of Pharmacology, Perelman School of Medicine, University of Pennsylvania, Philadelphia, PA, USA. eberwine@mail.med.upenn.edu.
authors.forename: James
authors.initials: J
authors.lastname: Eberwine
authors.affiliation: Department of Biology, School of Arts and Sciences, University of Pennsylvania, 301A/B Lynch Laboratory, 433 S University Avenue, Philadelphia, PA, 19104, USA. junhyong@sas.upenn.edu.
authors.forename: Junhyong
authors.initials: J
authors.lastname: Kim
mesh_terms: D000818:Animals; D002478:Cells, Cultured; D020869:Gene Expression Profiling; D014644:Genetic Variation; D059014:High-Throughput Nucleotide Sequencing; D051379:Mice; D008810:Mice, Inbred C57BL; D020871:RNA Stability; D051381:Rats; D017207:Rats, Sprague-Dawley; D017423:Sequence Analysis, RNA; D059010:Single-Cell Analysis; D059467:Transcriptome