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. 2023 Feb 16;2(1):e87.
doi: 10.1002/imt2.87. eCollection 2023 Feb.

Using PhyloSuite for molecular phylogeny and tree-based analyses

Chuan-Yu Xiang  1 , Fangluan Gao  2 , Ivan Jakovlić  1 , Hong-Peng Lei  1 , Ye Hu  1 , Hong Zhang  1 , Hong Zou  3 , Gui-Tang Wang  3 , Dong Zhang  1
Affiliations

Using PhyloSuite for molecular phylogeny and tree-based analyses

Chuan-Yu Xiang et al. Imeta. .

Abstract

Phylogenetic analysis has entered the genomics (multilocus) era. For less experienced researchers, conquering the large number of software programs required for a multilocus-based phylogenetic reconstruction can be somewhat daunting and time-consuming. PhyloSuite, a software with a user-friendly GUI, was designed to make this process more accessible by integrating multiple software programs needed for multilocus and single-gene phylogenies and further streamlining the whole process. In this protocol, we aim to explain how to conduct each step of the phylogenetic pipeline and tree-based analyses in PhyloSuite. We also present a new version of PhyloSuite (v1.2.3), wherein we fixed some bugs, made some optimizations, and introduced some new functions, including a number of tree-based analyses, such as signal-to-noise calculation, saturation analysis, spurious species identification, and etc. The step-by-step protocol includes background information (i.e., what the step does), reasons (i.e., why do the step), and operations (i.e., how to do it). This protocol will help researchers quick-start their way through the multilocus phylogenetic analysis, especially those interested in conducting organelle-based analyses.

Keywords: annotation; concatenation; iTOL; loci; multiple‐sequence alignment; partitioning; trimming.

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Conflict of interest statement

The authors declare no conflicts of interest.

Figures

Figure 1
Figure 1
Introduction to the PhyloSuite interface.
Figure 2
Figure 2
Downloading mitogenome data from the NCBI's Nucleotide database.
Figure 3
Figure 3
Creating a new work folder in PhyloSuite.
Figure 4
Figure 4
Importing the sequences into the PhyloSuite work folder.
Figure 5
Figure 5
Filtering redundant sequences.
Figure 6
Figure 6
Get taxonomy.
Figure 7
Figure 7
Extracting genes from mitochondrial genomes.
Figure 8
Figure 8
General settings and parameters for extraction.
Figure 9
Figure 9
Illustration of a multiple sequence alignment. Gaps are represented by “‐”.
Figure 10
Figure 10
Multiple sequence alignment using MAFFT.
Figure 11
Figure 11
Aligning customized sequences in MAFFT.
Figure 12
Figure 12
Optimization of multiple codon sequence alignments using MACSE.
Figure 13
Figure 13
Aligning customized sequences in MACSE.
Figure 14
Figure 14
Illustration of multiple sequence alignment trimming.
Figure 15
Figure 15
PCGs alignment trimming using Gblocks.
Figure 16
Figure 16
RNA alignment trimming using trimAl.
Figure 17
Figure 17
Trimming customized sequences using Gblocks.
Figure 18
Figure 18
Illustration of sequence concatenation.
Figure 19
Figure 19
Multiple sequence alignment concatenation of the PCGsRNA dataset.
Figure 20
Figure 20
Multiple sequence alignment concatenation of the PCGs12RNA dataset.
Figure 21
Figure 21
Multiple sequence alignment concatenation of AA.
Figure 22
Figure 22
Multiple sequence alignment concatenation of customized sequences.
Figure 23
Figure 23
Partitioning analysis using ModelFinder.
Figure 24
Figure 24
Partitioning and optimal model selection using PartitionFinder2.
Figure 25
Figure 25
Phylogenetic tree reconstruction using IQ‐TREE software.
Figure 26
Figure 26
Phylogenetic tree reconstruction using IQ‐TREE and customized input files.
Figure 27
Figure 27
Phylogenetic tree reconstruction using MrBayes in PhyloSuite.
Figure 28
Figure 28
Continuing previous analyses in MrBayes.
Figure 29
Figure 29
Phylogenetic tree reconstruction using MrBayes and customized input files.
Figure 30
Figure 30
The operation of TreeSuite.
Figure 31
Figure 31
Saturation analysis in TreeSuite.
Figure 32
Figure 32
Regression analysis of saturation plots. Here we used MrBayes results based on mitogenomes of 55 flatworms for illustration.
Figure 33
Figure 33
Calculation of the long‐branch score in TreeSuite.
Figure 34
Figure 34
Identification of spurious species in TreeSuite.
Figure 35
Figure 35
Treeness, RCV (relative composition variability) and signal‐to‐noise analyses in TreeSuite.
Figure 36
Figure 36
Pairwise patristic distance (branch length) and root‐to‐tip branch length calculation in TreeSuite.
Figure 37
Figure 37
Calculation of the evolution rate of multiple tree files in TreeSuite.
Figure 38
Figure 38
Unroot tree in TreeSuite.
Figure 39
Figure 39
Resolve polytomy in TreeSuite.
Figure 40
Figure 40
Mitochondrial genetic code for Gyrodactylidea.
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