Search research articles

ABOUT JoVE

Overview Leadership Blog JoVE Help Center

AUTHORS

Publishing Process Editorial Board Scope & Policies Peer Review FAQ Submit

LIBRARIANS

Testimonials Subscriptions Access Resources Library Advisory Board FAQ

RESEARCH

JoVE Journal Methods Collections JoVE Encyclopedia of Experiments Archive

EDUCATION

JoVE Core JoVE Business JoVE Science Education JoVE Lab Manual Faculty Resource Center Faculty Site

Terms & Conditions of Use

Search research articles

Related Experiment Video

Updated: May 17, 2026

An Allele-specific Gene Expression Assay to Test the Functional Basis of Genetic Associations

An Allele-specific Gene Expression Assay to Test the Functional Basis of Genetic Associations

Published on: November 3, 2010

Quantifying reference alignment bias in functional genomics analyses.

Nina Tekkey¹, John E Garza², Wenjin Zhang¹

¹Department of Genetics, Washington University School of Medicine, 660 South Euclid Ave., Saint Louis, MO, USA.

Cell Reports Methods

|May 15, 2026

Summary

Related Concept Videos

You might also read

Related Articles

Articles linked to this work by shared authors, journal, and citation graph.

Sort by

Same author

Genomic and physiological changes in a sexually selected and frugivorous bird radiation.

Current biology : CB·2026

Same author

Benchmarking genome choice in functional genomics analyses.

Nature communications·2026

Same author

Characterizing cytosine methylation of polymorphic transposable element insertions using the human pangenome resources.

Genome research·2026

Same author

Genealogy-based trait association with LOCATER boosts power at loci with allelic heterogeneity.

Genome research·2026

Same author

Fully T2T pedigree assemblies reveal genetic stability and epigenetic plasticity of human centromeres across inheritance and cell-fate transitions.

bioRxiv : the preprint server for biology·2026

Same author

Leveraging Human Pangenome for Improved Somatic Variant Detection.

bioRxiv : the preprint server for biology·2026

Same journal

A computational method to design broad-spectrum T cell-inducing vaccines applied to Betacoronaviruses.

Cell reports methods·2026

Same journal

MalDeepSeq panel: A targeted ultra-deep sequencing approach to trace drug resistance markers in Plasmodium falciparum.

Cell reports methods·2026

Same journal

Induced pluripotent stem cell-derived macrophages enable broad modeling of human inflammasome signaling.

Cell reports methods·2026

Same journal

Rapid discovery of cell-surface glycosylation regulators using a lectin-based magnetic CRISPR screen.

Cell reports methods·2026

Same journal

A real-time FRET ubiquitin transfer assay for quantitative characterization of ternary complexes in targeted protein degradation.

Cell reports methods·2026

Same journal

A high-throughput, end-to-end pipeline for extracellular miRNA biomarker discovery from human biofluids.

Cell reports methods·2026

See all related articles

This summary is machine-generated.

Reference alignment bias (RAB) systematically errors next-generation sequencing (NGS) data. This study quantifies RAB in functional genomics, revealing significant bias in RNA-seq, ATAC-seq, and WGBS, underscoring the need for representative reference genomes.

Area of Science:

Genomics
Bioinformatics
Computational Biology

Background:

Next-generation sequencing (NGS) alignment to a reference genome can introduce systematic errors, termed reference alignment bias (RAB).
The impact of RAB on functional genomics readouts remains incompletely quantified.
The Human Pangenome Reference Consortium provides valuable resources for studying genomic variation.

Purpose of the Study:

To quantify the extent of reference alignment bias (RAB) across different functional genomics assays.
To assess the impact of using the standard human reference genome (hg38) on data interpretation.
To highlight the importance of representative reference genomes in genomic studies.

Main Methods:

Leveraging data from the Human Pangenome Reference Consortium.

Keywords:

ATAC-seq CP: genetics RNA-seq WGBS functional genomics genetics genomics

More Related Videos

Optimization for Sequencing and Analysis of Degraded FFPE-RNA Samples

Optimization for Sequencing and Analysis of Degraded FFPE-RNA Samples

Published on: June 8, 2020

Related Experiment Videos

Last Updated: May 17, 2026

An Allele-specific Gene Expression Assay to Test the Functional Basis of Genetic Associations

An Allele-specific Gene Expression Assay to Test the Functional Basis of Genetic Associations

Published on: November 3, 2010

Optimization for Sequencing and Analysis of Degraded FFPE-RNA Samples

Optimization for Sequencing and Analysis of Degraded FFPE-RNA Samples

Published on: June 8, 2020

Quantifying RAB in RNA sequencing (RNA-seq), ATAC-seq, and whole-genome bisulfite sequencing (WGBS) assays.

Analyzing alignment data against the hg38 human reference genome.

Main Results:

On average, 0.2% of the genome is susceptible to RAB in RNA-seq studies.
Approximately 1% of the genome shows bias in ATAC-seq.
Up to 3% of the genome is affected by RAB in WGBS studies using the hg38 reference.

Conclusions:

Reference alignment bias (RAB) significantly impacts functional genomics assays.
The choice of reference genome critically influences the accuracy of genomic data interpretation.
Utilizing more representative reference genomes is crucial for mitigating RAB and improving functional genomics studies.