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Updated: Nov 5, 2025

Microbiota Analysis Using Two-step PCR and Next-generation 16S rRNA Gene Sequencing
Published on: October 15, 2019
Identifying biases and their potential solutions in human microbiome studies
Jacob T Nearing1, André M Comeau2, Morgan G I Langille3,4
1Department of Microbiology and Immunology, Dalhousie University, Halifax, Nova Scotia, Canada.
Abstract:
Advances in DNA sequencing technology have vastly improved the ability of researchers to explore the microbial inhabitants of the human body. Unfortunately, while these studies have uncovered the importance of these microbial communities to our health, they often do not result in similar findings. One possible reason for the disagreement in these results is due to the multitude of systemic biases that are introduced during sequence-based microbiome studies. These biases begin with sample collection and continue to be introduced throughout the entire experiment leading to an observed community that is significantly altered from the true underlying microbial composition. In this review, we will highlight the various steps in typical sequence-based human microbiome studies where significant bias can be introduced, and we will review the current efforts within the field that aim to reduce the impact of these biases. Video abstract.
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