Please see the documentation website for more details.
MitoPilot is a package for the assembly and annotation of mitochondrial genomes from genome skimming data. The core application consists of a Nextflow pipeline that is wrapped in an R package, which includes an R-Shiny graphical interface to monitor and interact with processing parameters and outputs. Currently the pipeline expects paired-end Illumina reads as the raw input and performs the following steps.
- Mitogenome assembly
- fastp for quality control and adapter trimming
- GetOrganelle (default) or MitoFinder for mitogenome assembly
- bowtie2 for read mapping to calculate coverage and error rates.
- NCBI BLAST against a local database of all annotated metazoan mitogenomes in GenBank, packaged in the MitoPilot container, to find the closest reference for automatic and manual curation
- Mitogenome annotation
- MITOS2 for rRNA, PCG, and tRNA annotation
- tRNAscan-SE for tRNA annotation
- MitoFinder for rRNA and PCG annotation (optional)
- ARWEN for tRNA annotation (optional)
- ARAGORN for tRNA annotation (optional)
- ORFfinder identify additional open reading frames (ORFs) (optional)
- Custom scripts for gene boundary refinement and annotation file formatting
- Validation to flag possible issues or known errors that would be rejected by NCBI GenBank
- Manual curation of annotations using the integrated Shiny App
- Data export
- Custom scripts to export data in a format suitable for submission to NCBI GenBank
Optionally, MitoPilot can proceed straight to annotation and curation if
the user supplies mitogenome assemblies with the
new_project_userAsmb() function.
MitoPilot needs R (>= 4.4.0), Java 17+, Nextflow (24.10.x - 25.10.x), and a container runtime (Docker locally, or Singularity/Apptainer on a cluster).
if (!requireNamespace("BiocManager", quietly = TRUE)) {
install.packages("BiocManager")
}
BiocManager::install("Smithsonian/MitoPilot")See Installation and Requirements for the full requirements, disk space, updating and version pinning, and container cache setup. Cluster-specific instructions are available for Smithsonian Hydra and NOAA SEDNA; for any other cluster see HPC cluster support.
MitoPilot ships a small pre-filtered test dataset. Running it end to end is the recommended way to verify your installation and learn the interface before using your own data.
The Get Started tutorial walks the whole pipeline using this test project.
Want to skip straight to using MitoPilot with your own data? Head on over to Starting Your Own Project.
MitoPilot was initially built for fish mitogenomes, but It has since been extended with curation and validation rulesets for the groups below.
| Clade | Common name | curate_target | Status |
|---|---|---|---|
| Actinopterygii | Ray-finned fishes | fish_mito | Tested |
| Annelida | Annelids | annelid_mito | Testing in progress |
| Ascidiacea | Sea squirts | ascidiacea_mito | Untested |
| Asteroidea | Sea stars | starfish_mito | Tested |
| Aves | Birds | bird_mito | Untested |
| Bivalvia | Bivalves | bivalvia_mito | Untested |
| Bryozoa | Bryozoans | bryozoa_mito | Untested |
| Copepoda | Copepods | copepod_mito | Testing in progress |
| Crinoidea | Crinoids | crinoidea_mito | Untested |
| Ctenophora | Ctenophores | ctenophore_mito | Testing in progress |
| Demospongiae | Demosponges | demospongiae_mito | Untested |
| Diptera | True flies | diptera_mito | Tested |
| Echinoidea | Sea urchins | echinoidea_mito | Untested |
| Gastropoda | Gastropods | gastropoda_mito | Testing in progress |
| Hexacorallia | Hexacorals | hexacoral_mito | Tested |
| Holothuroidea | Sea cucumbers | holothuroidea_mito | Untested |
| Homoscleromorpha | Homoscleromorph sponges | homoscleromorpha_mito | Untested |
| Hydrozoa | Hydrozoans | hydrozoa_mito | Testing in progress |
| Lepidosauria | Lepidosaurs | lepidosaur_mito | Untested |
| Malacostraca | Malacostracans | malacostraca_mito | Testing in progress |
| Mammalia | Mammals | mammal_mito | Untested |
| Nemertea | Ribbon worms | nemertea_mito | Testing in progress |
| Octocorallia | Octocorals | octocoral_mito | Tested |
| Ophiuroidea | Brittle stars | ophiuroidea_mito | Untested |
| Platyhelminthes | Flatworms | platyhelminthes_mito | Untested |
| Polychaeta | Polychaetes | polychaeta_mito | Testing in progress |
| Pycnogonida | Sea spiders | pycnogonida_mito | Untested |
| Scyphozoa | True jellyfishes | scyphozoa_mito | Testing in progress |
| Sipuncula | Peanut worms | sipuncula_mito | Untested |
| Testudines | Turtles | turtle_mito | Tested |
| Thaliacea | Salps | thaliacea_mito | Testing in progress |
| Thecostraca | Barnacles | thecostraca_mito | Untested |
See the curation ruleset
browser
for details about each curation ruleset. The curation ruleset can be set
individually for each sample in the Curate Opts. window in the
MitoPilot app.
All curation rulesets ship inside the Docker image (macguigand/MitoPilot). If MitoPilot doesn’t have a curation ruleset for your taxonomic group, please open an issue or contact Dan MacGuigan at macguigand@si.edu.
For groups other than fishes, make sure you build or pick the appropriate reference databases. There are three independent kinds of databases:
- Assembly references for GetOrganelle or MitoFinder.
MitoPilot::custom_assembly_db()builds these for a clade automatically, with no external tools required. See building custom databases. - Annotation references for MITOS2. MitoPilot includes Chordata
and Metazoa databases, selectable in the
Annotate Opts.window. - Curation references, chosen independently of the annotation
database. Bundled options are
Metazoa_RefSeq235(the default),Metazoa_RefSeq231,Metazoa_RefSeq89, andChordata. MitoPilot also folds each sample’s assembly BLAST results into the curation references automatically.
| Page | What it covers |
|---|---|
| Get Started | Full walkthrough, then starting your own project |
| Installation and Requirements | Prerequisites, installing, updating, container cache |
| HPC cluster support | Executors, cluster profiles, SSH tunnel, submitting runs |
| Curation ruleset browser | What each clade ruleset enforces |
| Building custom databases | Assembly and curation reference databases |
| Handling difficult assemblies | Resolving competing assemblies and fragmented scaffolds |
| FAQ and Troubleshooting | Common questions and pipeline failures |
| Reference | All functions |
| Changelog | Release notes and container tags |