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next-generation; sequencing; Escherichia coli; Pseudomonas syringae
Abstract:
Studies of microbial evolutionary dynamics are being transformed by the availability of affordable high-throughput
sequencing technologies, which allow whole-genome sequencing of hundreds of related taxa in a single study.
Reconstructing a phylogenetic tree of these taxa is generally a crucial step in any evolutionary analysis. Instead of
constructing genome assemblies for all taxa, annotating these assemblies, and aligning orthologous genes, many
recent studies 1) directly map raw sequencing reads to a single reference sequence, 2) extract single nucleotide polymorphisms
(SNPs), and 3) infer the phylogenetic tree using maximumlikelihood methods from the aligned SNP positions.
However, here we show that, when using such methods to reconstruct phylogenies from sets of simulated sequences, both
the exclusion of nonpolymorphic positions and the alignment to a single reference genome, introduce systematic biases
and errors in phylogeny reconstruction. To address these problems, we developed a new method that combines
alignments from mappings to multiple reference sequences and show that this successfully removes biases from the
reconstructed phylogenies. We implemented this method as a web server named REALPHY (Reference sequence
Alignment-based Phylogeny builder), which fully automates phylogenetic reconstruction from raw sequencing reads.
