ArticleNAR genomics and bioinformatics2022
Internal oligo(dT) priming introduces systematic bias in bulk and single-cell RNA sequencing count data.
Article in NAR genomics and bioinformatics, 2022. The graph could read no effect estimate from its abstract, so it casts no vote on the map. Cited by 34 papers.
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Who cites it
34 citing papers in PubMed, 39 citations in OpenAlex.
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- Increasing usable reads in RNA-seq protocols.iScience · 2026Article
- scPASU: A computational protocol for quantifying polyadenylation site usage and alternative polyadenylation from 3' scRNA-seq data.STAR protocols · 2026Article
- StringTie3 improves total RNA-seq assembly by resolving nascent and mature transcripts.Nature methods · 2026Article
- Optimizing Single-Cell Long-Read Sequencing for Enhanced Isoform Detection in Pancreatic Islets.Diabetes · 2026Article
- A practical guide to targeted single-cell RNA sequencing technologies.Communications biology · 2026Review
- PolyAseqTrap: a universal tool for genome-wide identification and quantification of polyadenylation sites from different 3' end sequencing data.Genome biology · 2026Article
- Differentiation in the human urothelia is defined by distinct alternative polyadenylation.Cell reports · 2026Article
- PolyA_DB v4: systematic polyA site identification and isoform annotation in human and mouse genomes using 3' end and long-read sequencing data.Nucleic acids research · 2026Article
- Multisite Assessment of Methods for Cell Preservation Upstream of Single-Cell RNA Sequencing.Journal of biomolecular techniques : JBT · 2026Article
- Biomaterial-mediated Cell Atlas: an insight from single-cell and spatial transcriptomics.Bioactive materials · 2025Review
- Multi-site Assessment of Methods for Cell Preservation Upstream of Single Cell RNA Sequencing.bioRxiv : the preprint server for biology · 2025Article
- Long-read RNA sequencing dataset of human pancreatic cancer cell lines.Scientific data · 2025Article
- Optimizing Single-Cell Long-Read Sequencing for Enhanced Isoform Detection in Pancreatic Islets.bioRxiv : the preprint server for biology · 2025Article
- Article
- Computational methods for alternative polyadenylation and splicing in post-transcriptional gene regulation.Experimental & molecular medicine · 2025Review
- Spatial-scERA: a method for reconstructing spatial single-cell enhancer activity in multicellular organisms.Nucleic acids research · 2025Article
- Efficient profiling of total RNA in single cells with STORM-seq.bioRxiv : the preprint server for biology · 2025Article
- MSI expresso: a software for determining MSI status and detecting MSI-related transcription events from RNA sequencing data.Frontiers in genetics · 2025Article
Corrections and comments
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Authors and funding
3 authors at 1 institution in 1 country.
Funding
Abstract
Significant advances in RNA sequencing have been recently made possible by using oligo(dT) primers for simultaneous mRNA enrichment and reverse transcription priming. The associated increase in efficiency has enabled more economical bulk RNA sequencing methods and the advent of high-throughput single-cell RNA sequencing, already one of the most widely adopted methods in transcriptomics. However, the effects of off-target oligo(dT) priming on gene expression quantification have not been appreciated. In the present study, we describe the extent, the possible causes, and the consequences of internal oligo(dT) priming across multiple public datasets obtained from various bulk and single-cell RNA sequencing platforms. To explore and address this issue, we developed a computational algorithm for RNA counting methods, which identifies the sequencing read alignments that likely resulted from internal oligo(dT) priming and removes them from the data. Directly comparing filtered datasets to those obtained by an alternative method reveals significant improvements in gene expression measurement. Finally, we infer a list of human genes whose expression quantification is most likely to be affected by internal oligo(dT) priming and predict that when measured using these methods, the expression of most genes may be inflated by at least 10% whereby some genes are affected more than others.
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What OpenQuestion holds
Registered trials
Read under generation 80e0d062 · epoch 390. Bibliography from PubMed, PubMed Central and OpenAlex; grants from NIH RePORTER; trial links from ClinicalTrials.gov; estimates, votes and beliefs from the OpenQuestion graph.