phylogeny.fr

2.1 "One Click" mode

This is the "default" mode which proposes a pipeline already set up to run and connect programs recognized for their accuracy and speed to reconstruct a robust phylogenetic tree from a set of sequences:

• MAFFT has been chosen for rapid and robust multiple sequence alignment.
• ClipKit can optionally be used for alignment curation (trimming unreliable regions).
• FastTree (Fast mode) or IQ-TREE (Accurate mode) can be selected for phylogeny.

You have the ability to toggle the curation step (ClipKit) to eliminate poorly aligned positions and highly divergent regions before the tree inference step. At the end of the analysis, an interactive and exportable rendition of the generated tree will be available.

Default Parameters

For transparency, below is the exhaustive list of the parameters used by the tools behind the scenes during a One Click run:

2.2 Advanced mode

The Advanced mode follows the same robust pipeline philosophy as "One Click" mode, utilizing a predefined succession of programs (MAFFT, ClipKit, and IQ-TREE). However, it provides you with complete control over the parameters for each step of the analysis.

2.3 "A la carte" mode

The "A la carte" mode provides the ultimate flexibility for building your phylogenetic analysis pipeline. This mode allows you to handpick a tool for each step of the analysis, giving complete control over the methodology.

Two-Phase Workflow

The A la carte mode operates in two distinct phases:

1. Pipeline Design: Select the tools to include in the analysis. A visual pipeline at the top of the page shows the current selection in real-time.
2. Configuration & Launch: After confirming the tool selection, configure the settings for each tool individually, upload the input FASTA file, and launch the analysis.

How It Works

1. Navigate to the A la carte page from the main menu.
2. Click on the tool tiles to select your preferred tool for each step (Alignment, Curation, Phylogeny). Selected tools are highlighted with a checkmark.
3. Click "Continue to Configuration" to proceed.
4. Upload your FASTA file or paste your sequences directly.
5. Configure each selected tool's parameters using the provided forms.
6. Optionally enter your email to receive a notification when the analysis completes.
7. Click "Submit Analysis" to start the pipeline.

3.1 Submitting a query

To run a BLAST search, a single sequence in FASTA format has to be provided either as a file or by copying and pasting it directly into the text box.

BLAST Programs

Users have to select the appropriate program based on the query and target database types:

Available Databases

A variety of sequence databases are available:

E-value Threshold

The E-value (Expect value) represents the number of hits expected by chance. A lower E-value indicates a more significant match:

• 1.e-5 (default): Good balance between sensitivity and specificity.
• 1.e-10 to 1.e-30: More stringent, returns only highly confident hits.
• 0.1 to 1: More permissive, may include distant homologs.

Optional Clustering

MMseqs2 can be used to cluster all results. For instance, as the number of returned sequences is limited, this can avoid the over-representation of one kind of organism.

3.2 Interactive results

Once the BLAST job completes, an interactive visualization is displayed, combining a phylogenetic tree with a floating Toolbox panel dedicated to filtering and exporting results.

Phylogenetic Tree Display

The tree displays the query sequence alongside the top BLAST hits, making it possible to visualize evolutionary relationships at a glance:

• Query highlighting: The input sequence is visually emphasized in the tree.
• Taxonomy coloring: Sequences are colored according to taxonomic groups (with adjustable depth).
• Interactive selection: Tree leaves can be clicked directly to select or deselect sequences.
• External links: Sequence IDs provide access to the corresponding original database entries.

The Toolbox Panel

The floating Toolbox serves as the main control center for exploring BLAST results. It can be moved freely on the screen and provides two distinct modes accessible through tabs.

Tree View Tab

This tab focuses on the top hits displayed in the phylogenetic tree (typically the highest-scoring sequences). It is intended for precise, small-scale selection:

• Selection counter: Displays the number of currently selected sequences relative to the total number of tree leaves.
• Quick select buttons: Allows all tree sequences to be selected or deselected instantly.
• Bitscore filter pills: Filters sequences according to score ranges (<40, 40–50, 50–80, 80–200, >200) using color-coded filters.
• Taxonomy depth slider: Adjusts the displayed taxonomic classification level.
• Gap-free indicator: Displays the percentage of alignment positions without gaps for the current selection, providing an indicator of alignment quality.

All Hits Tab

This tab operates on all BLAST hits, limited to 500, not only those displayed in the tree. It is intended for large-scale filtering across the complete result set:

• E-value threshold: Dynamically filters hits according to E-value (from 1e-5 to 1e-100).
• Histogram filters: Interactive histograms allow sequence selection based on:
  • Score (bitscore): Filtering by alignment score strength.
  • Similarity (% identity): Filtering by sequence similarity percentage.
  • Coverage (% query coverage): Filtering by query coverage percentage.
• Taxonomy filter: A hierarchical taxonomy tree enables selection of sequences belonging to specific taxonomic groups (for example, only Bacteria or only Vertebrates).
• Select in table: A selection tool based on the BLAST table results.
• Reset selection: Clears all filters and restores the initial state.

Exporting & Pipeline Integration

Both tabs provide export options for the selected sequences:

• Download FASTA: Export selected sequences as a FASTA file for external analysis.
• Send to Pipeline: Directly transfer selected sequences to the One Click or Advanced phylogeny pipelines. The sequences are automatically loaded into the analysis form.

Taxonomy Legend

Below the tree, a dynamic legend displays the taxonomic groups present in the results with their associated colors. The groups shown depend on the taxonomy depth setting in the Toolbox.