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Power analysis of single-cell RNA-sequencing experiments

Accepted version
Peer-reviewed

Type

Article

Change log

Authors

Svensson, V 
Natarajan, KN 
Ly, L-H 
Miragaia, RJ 
Labalette, C 

Abstract

Single-cell RNA sequencing (scRNA-seq) has become an established and powerful method to investigate transcriptomic cell-to-cell variation, thereby revealing new cell types and providing insights into developmental processes and transcriptional stochasticity. A key question is how the variety of available protocols compare in terms of their ability to detect and accurately quantify gene expression. Here, we assessed the protocol sensitivity and accuracy of many published data sets, on the basis of spike-in standards and uniform data processing. For our workflow, we developed a flexible tool for counting the number of unique molecular identifiers (https://github.com/vals/umis/). We compared 15 protocols computationally and 4 protocols experimentally for batch-matched cell populations, in addition to investigating the effects of spike-in molecular degradation. Our analysis provides an integrated framework for comparing scRNA-seq protocols.

Description

Keywords

Animals, Embryonic Stem Cells, Freezing, Mice, Poly A, RNA, Messenger, Sensitivity and Specificity, Sequence Analysis, RNA, Single-Cell Analysis, Workflow

Journal Title

Nature Methods

Conference Name

Journal ISSN

1548-7091
1548-7105

Volume Title

14

Publisher

Nature Publishing Group
Sponsorship
Medical Research Council (MC_PC_12009)
European Research Council (677501)
The study was supported by Cancer Research UK grant number C45041/A14953 to A Cvejic and C Labalette, European Research Council project 677501-ZF_Blood to A Cvejic and a core support grant from the Wellcome Trust and MRC to the Wellcome Trust–Medical Research Council Cambridge Stem Cell Institute. The ERC grant ThSWITCH to SA Teichmann (grant no. 260507) and a Lister Institute Research Prize to SA Teichmann. KN Natarajan was supported by the Wellcome Trust Strategic Award “Single cell ge nomics of mouse gastrulation”.