Taxonomy-guided multiple sequence alignment for difficult genes, including 12S and 16S mitochondrial rRNA.
Use to perform multiple sequence alignment for a highly variable gene and a large number of divergent sequences.
TaxonomyAlign automatically divides the sequences into smaller subsets based on user-supplied taxonomic groupings, and builds sub-alignments for the groups using MAFFT (several algorithm options available).
The defined groupings should be related to both taxonomy and phylogeny (e.g., genus, family, superfamily, etc).
The resulting sub-alignments are then merged together with MAFFT, with or without an optional polishing step. If a grouping only contains a single sequence, that sequence will be added to the alignments during the merge step.
The original use-case was for creating high-quality alignments from many 12S and 16S mtDNA sequences across a large phylogenetic scale. However, TaxonomyAlign should prove useful for other genes with many species and divergent sequences to align.
The current release of TaxonomyAlign is v1.0.
TaxonomyAlign is written in Python (compatible with 3.7) and functions as a command-line script (TaxonomyAlign.py). It can be downloaded and executed independently without the need to install TaxonomyAlign as a Python package or library.
TaxonomyAlign requires MAFFT and BioPython (v1.77).
Installation of these requirements is fast and easy using conda. The taxonomyalign-conda-env.yml file can be used to create the correct conda environment:
conda create -f taxonomyalign-conda-env.yml
The resulting conda environment can then be activated using:
conda activate taxonomyalign
You can then run TaxonomyAlign in this environment.
TaxonomyAlign was introduced in the following publication:
Portik, D.M., Streicher, J.W., and J.J. Wiens. 2023. Frog phylogeny: a time-calibrated, species-level tree based on hundreds of loci and 5,242 species. Molecular Phylogenetics and Evolution, 188: 107907.
Please cite this work if you are using TaxonomyAlign for a publication.
TaxonomyAlign provides an alternative to other alignment methods. It uses taxonomy-guided multiple sequence alignment for difficult genes. It was designed for large-scale 12S and 16S rRNA alignments for vertebrates (>3,000 sequences). However, it should prove useful for other genes with many species and divergent sequences to align.
Example datasets are provided in the example data folder. There are two fasta files containing either 12S or 16S sequences, for all frogs. The map file (Anura-Family-Map-File.txt) can be used with either file to run the analysis.
This is a tab-delimited text file with two columns. The first column should contain the name of the taxon/sequence. The second column should contain the taxonomic group name that the taxon is a part of. This could be genus, family, or higher rank. For frogs, I used family level groupings. An example of this is shown below, and the full file can be found in the example data folder.
Semnodactylus_wealii Hyperoliidae
Tachycnemis_seychellensis Hyperoliidae
Leiopelma_archeyi Leiopelmatidae
Leiopelma_hamiltoni Leiopelmatidae
Leiopelma_hochstetteri Leiopelmatidae
Leiopelma_pakeka Leiopelmatidae
Adenomera_ajurauna Leptodactylidae
Adenomera_andreae Leptodactylidae
Adenomera_araucaria Leptodactylidae
You can use any method to generate the map file, but it is important to ensure all sequences are assigned. Any sequence not appearing in this file will be excluded from the alignment. The map file can contain species that do not appear in your fasta file.
There is a quick way to obtain all of the sequence names from your unaligned fasta file using the --output_seqnames flag:
python TaxonomyAlign.py -f <fasta file> -m <map file> -o <output directory> --output_seqnames
The output from this option is a text file of the sequence names. This can be used to create the map file, but you will need to add the groupings.
Once the map file has been created, the analysis can be run. The general usage is shown below:
python TaxonomyAlign.py -f <fasta file> -m <map file> -o <output directory> -a <alignment algorithm> -t <threads>
The full path to an unaligned fasta file that contains DNA sequences.
The map file described in the above section.
The full path to an existing directory to write the output files.
The alignment algorithm to use. Choices include auto, FFT-NS-i, E-INS-i, L-INS-i, and G-INS-i. Defaults to auto. auto: auto-select based on input sequences FFT-NS-i: a progressive and general algorithm E-INS-i: best for several conserved core regions and variable regions (e.g., 12S and 16S mtDNA) L-INS-i: best for a single conserved core region with variable flanking regions (e.g., UCEs) G-INS-i: best for full-length single core sequences
Specifies number of threads to use.
python TaxonomyAlign.py -f bin/Analysis/12S-Anura.fasta -m bin/Analysis/Anura-Family-Map-File.txt -o bin/Analysis/Output1/ -a E-INS-i -t 8
The above command will align the fasta file 12S-Anura.fasta using the E-INS-i algorithm, creating sub-alignments from the map file Anura-Family-Map-File.txt. It will use 8 threads.
Performs additional distance measure refinement on final merged alignment, at cost of increasing run time (sometimes substantially).
Show mafft progress during final merge step instead of redirecting to log file. May be useful for monitoring the --polish option, which can be slow for larger datasets
Write output file of sequence names and quit. See above section for usage.
There are three output directories created. Each contains files unique to particular steps of the analysis. An example of the output directory structure is shown below:
output_dir/
│
├── 1-Group-Fasta-Files/
│ ├── Alytidae.multiples.fasta
│ ├── Arthroleptidae.multiples.fasta
│ ├── Ascaphidae.singleton.fasta
│ └── Batrachylidae.multiples.fasta
│
├── 2-Group-Alignments/
│ ├── Alytidae.mafft-E-INS-i.fasta
│ ├── Alytidae.mafft-E-INS-i.log
│ ├── Arthroleptidae.mafft-E-INS-i.fasta
│ ├── Arthroleptidae.mafft-E-INS-i.log
│ ├── Ascaphidae.mafft-E-INS-i.fasta
│ ├── Ascaphidae.mafft-E-INS-i.log
│ ├── Batrachylidae.mafft-E-INS-i.fasta
│ └── Batrachylidae.mafft-E-INS-i.log
│
├── 3-Merged-Alignments/
│ ├── Merged-alignment.fasta
│ ├── Merge-Table.txt
│ ├── Merged-alignment.log
│ └── Sub-Alignments.fasta
│
└── Taxonomy-Align_Aug-06-2022_12.16.14.log
1-Group-Fasta-Files/
This directory will contain fasta files for each grouping included in the map file. The multiples or singleton component of the name signify whether the file contains >= 2 sequences (multiples) or a single sequence (singleton). The multiples files are individually aligned in the next step. The singleton files only contain one sequence and they are aligned during the final sub-alignment merging step.
2-Group-Alignments/
This directory contains the aligned outputs of the group-specific fasta files. The associated log files show the MAFFT progress.
3-Merged-Alignments/
This directory contains the final outputs.
-
Merged-alignment.fasta: This is the final alignment produced by TaxonomyAlign, and the main output file of interest. -
Merge-Table.txt: This MAFFT-specific file contains information about how the group merging was conducted. It is used in conjunction with theSub-Alignments.fastafile. -
Merged-alignment.log: Contains all of the MAFFT alignment progress reports. If using--verbose, this will also appear on the screen. -
Sub-Alignments.fasta: A concatenated sub-alignments fasta file that is specific to MAFFT. It is used in conjunction with theMerge-Table.txtfile.
GNU Lesser General Public License v3.0
TaxonomyAlign is written and maintained by Daniel Portik.
If you experience any problems running TaxonomyAlign, or have questions, please open an issue here.