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      The Human Cell Atlas: Technical approaches and challenges

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          Abstract

          The Human Cell Atlas is a large, international consortium that aims to identify and describe every cell type in the human body. The comprehensive cellular maps that arise from this ambitious effort have the potential to transform many aspects of fundamental biology and clinical practice. Here, we discuss the technical approaches that could be used today to generate such a resource and also the technical challenges that will be encountered.

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          Comparative Analysis of Single-Cell RNA Sequencing Methods.

          Christoph Ziegenhain, Beate Vieth, Swati Parekh … (2017)
          Single-cell RNA sequencing (scRNA-seq) offers new possibilities to address biological and medical questions. However, systematic comparisons of the performance of diverse scRNA-seq protocols are lacking. We generated data from 583 mouse embryonic stem cells to evaluate six prominent scRNA-seq methods: CEL-seq2, Drop-seq, MARS-seq, SCRB-seq, Smart-seq, and Smart-seq2. While Smart-seq2 detected the most genes per cell and across cells, CEL-seq2, Drop-seq, MARS-seq, and SCRB-seq quantified mRNA levels with less amplification noise due to the use of unique molecular identifiers (UMIs). Power simulations at different sequencing depths showed that Drop-seq is more cost-efficient for transcriptome quantification of large numbers of cells, while MARS-seq, SCRB-seq, and Smart-seq2 are more efficient when analyzing fewer cells. Our quantitative comparison offers the basis for an informed choice among six prominent scRNA-seq methods, and it provides a framework for benchmarking further improvements of scRNA-seq protocols.
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            Quantitative single-cell RNA-seq with unique molecular identifiers.

            Saiful Islam, Amit Zeisel, Simon Joost … (2014)
            Single-cell RNA sequencing (RNA-seq) is a powerful tool to reveal cellular heterogeneity, discover new cell types and characterize tumor microevolution. However, losses in cDNA synthesis and bias in cDNA amplification lead to severe quantitative errors. We show that molecular labels--random sequences that label individual molecules--can nearly eliminate amplification noise, and that microfluidic sample preparation and optimized reagents produce a fivefold improvement in mRNA capture efficiency.
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              Genetic programs in human and mouse early embryos revealed by single-cell RNA sequencing.

              Zhigang Xue, Kevin Huang, Chaochao Cai … (2013)
              Mammalian pre-implantation development is a complex process involving dramatic changes in the transcriptional architecture. We report here a comprehensive analysis of transcriptome dynamics from oocyte to morula in both human and mouse embryos, using single-cell RNA sequencing. Based on single-nucleotide variants in human blastomere messenger RNAs and paternal-specific single-nucleotide polymorphisms, we identify novel stage-specific monoallelic expression patterns for a significant portion of polymorphic gene transcripts (25 to 53%). By weighted gene co-expression network analysis, we find that each developmental stage can be delineated concisely by a small number of functional modules of co-expressed genes. This result indicates a sequential order of transcriptional changes in pathways of cell cycle, gene regulation, translation and metabolism, acting in a step-wise fashion from cleavage to morula. Cross-species comparisons with mouse pre-implantation embryos reveal that the majority of human stage-specific modules (7 out of 9) are notably preserved, but developmental specificity and timing differ between human and mouse. Furthermore, we identify conserved key members (or hub genes) of the human and mouse networks. These genes represent novel candidates that are likely to be key in driving mammalian pre-implantation development. Together, the results provide a valuable resource to dissect gene regulatory mechanisms underlying progressive development of early mammalian embryos.
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                Author and article information

                Journal
                Journal ID (nlm-ta): Brief Funct Genomics
                Journal ID (iso-abbrev): Brief Funct Genomics
                Journal ID (publisher-id): bfgp
                Title: Briefings in Functional Genomics
                Publisher: Oxford University Press
                ISSN (Print): 2041-2649
                ISSN (Electronic): 2041-2657
                Publication date Collection: July 2018
                Publication date (Electronic): 28 October 2017
                Publication date PMC-release: 28 October 2017
                Volume: 17
                Issue: 4 , Special Issue: Single-cell genomics
                Pages: 283-294
                Affiliations
                [1 ]RIKEN Center for Life Science Technologies, Division of Genomic Technologies, Yokohama, Kanagawa, Japan
                [2 ]Wellcome Trust Sanger Institute, Wellcome Genome Campus, Hinxton, Cambridge, UK
                Author notes
                Corresponding author: Michael J.T. Stubbington, Wellcome Trust Sanger Institute, Wellcome Genome Campus, Hinxton, Cambridge, CB10 1SA, UK. E-mail: ms31@ 123456sanger.ac.uk
                Author information
                http://orcid.org/0000-0001-5924-3566
                Article
                Publisher ID: elx029
                DOI: 10.1093/bfgp/elx029
                PMC ID: 6063304
                PubMed ID: 29092000
                SO-VID: b2fab478-ffa7-494a-a8aa-557f4f58cbba
                Copyright © © The Author 2017. Published by Oxford University Press.
                License:

                This is an Open Access article distributed under the terms of the Creative Commons Attribution License ( http://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.

                History
                Page count
                Pages: 12
                Funding
                Funded by: MEXT 10.13039/501100001700
                Funded by: RIKEN Center for Life Science Technologies
                Funded by: Wellcome Trust 10.13039/100004440
                Award ID: 206194
                Categories
                Subject: Papers

                ScienceOpen disciplines: Genetics
                Keywords: human cell atlas,single cell,rna sequencing,bioinformatics
                Data availability:
                ScienceOpen disciplines: Genetics
                Keywords: human cell atlas, single cell, rna sequencing, bioinformatics

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