BUTTERFLY: addressing the pooled amplification paradox with unique molecular identifiers in single-cell RNA-seq

BUTTERFLY: addressing the pooled amplification paradox with unique molecular identifiers in single-cell RNA-seq

Abstract The incorporation of unique molecular identifiers (UMIs) in single-cell RNA-seq assays makes possible the identification of duplicated molecules, thereby facilitating the counting of distinct molecules from sequenced reads. However, we show that the naïve removal of duplicates can lead to a...

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Bibliographic Details
Main Authors: Johan Gustafsson, Jonathan Robinson, Jens Nielsen, Lior Pachter
Format: Article
Language:English
Published: BMC 2021-06-01
Series:Genome Biology
Subjects:
Single-cell RNA-Seq
UMI
Droplet-based
PCR
Bias
Amplification
Online Access:https://doi.org/10.1186/s13059-021-02386-z
Description
Summary:Abstract The incorporation of unique molecular identifiers (UMIs) in single-cell RNA-seq assays makes possible the identification of duplicated molecules, thereby facilitating the counting of distinct molecules from sequenced reads. However, we show that the naïve removal of duplicates can lead to a bias due to a “pooled amplification paradox,” and we propose an improved quantification method based on unseen species modeling. Our correction called BUTTERFLY uses a zero truncated negative binomial estimator implemented in the kallisto bustools workflow. We demonstrate its efficacy across cell types and genes and show that in some cases it can invert the relative abundance of genes.
ISSN:1474-760X

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