Current RNA-seq methodology reporting limits reproducibility.

Area of Science:

• Bioinformatics
• Genomics
• Molecular Biology

Background:

• Ribonucleic acid sequencing (RNA-seq) is a key technology for quantifying RNA molecules.
• Bioinformatics pipelines are essential for processing raw RNA-seq data into gene or isoform counts.
• Current RNA-seq studies often overlook biases introduced by pipeline choices and biological references.

Purpose of the Study:

• To highlight the impact of methodological information on RNA-seq study reproducibility.
• To demonstrate how current practices prevent reliable replication of RNA-seq experiments.
• To advocate for standardized reporting of bioinformatics methods in RNA-seq.

Main Methods:

• Analysis of common practices in RNA-seq bioinformatics pipelines.
• Identification of sources of bias in RNA-seq data analysis.
• Review of methodological reporting standards in peer-reviewed RNA-seq literature.

Main Results:

• Failure to specify crucial methodological details in RNA-seq studies compromises reproducibility.
• Pipeline and biological reference selection introduce significant, often unacknowledged, biases.
• Existing reporting standards are insufficient for ensuring consistent RNA-seq experimental outcomes.

Conclusions:

• Explicit and standardized display of methodological information is necessary for RNA-seq reproducibility.
• Addressing pipeline and reference biases requires transparent reporting.
• Adoption of standardized reporting will enhance the reliability and comparability of RNA-seq research.