Abstract
As next-generation sequencing technologies advance rapidly and the cost of metagenomic sequencing continues to decrease, researchers now face an unprecedented volume of microbiome data. This surge has stimulated the development of scalable microbiome data analysis methods and necessitated the incorporation of phylogenetic information into microbiome analysis for improved accuracy. Tools for constructing phylogenetic trees from 16S rRNA sequencing data are well-established, as the highly conserved regions of the 16S gene are limited, simplifying the identification of marker genes. In contrast, metagenomic and whole genome shotgun (WGS) sequencing involve sequencing from random fragments of the entire gene, making identification of consistent marker genes challenging owing to the vast diversity of genomic regions, resulting in a scarcity of robust tools for constructing phylogenetic trees. Although bacterial sequence tree construction tools exist for upstream bioinformatics, many downstream researchers—those integrating these trees into statistical models or machine learning—are either unaware of these tools or find them difficult to use due to the steep learning curve of processing raw sequences. This is compounded by the fact that public datasets often lack phylogenetic trees, providing only abundance tables and taxonomic classifications. To address this, we present a comprehensive review of phylogenetic tree construction techniques for microbiome data (16S rRNA or whole-genome shotgun sequencing). We outline the strengths and limitations of current methods, offering expert insights and step-by-step guidance to make these tools more accessible and widely applicable in quantitative microbiome data analysis.