EAGLER is a scaffolding tool for long reads. The scaffolder takes as input a draft genome created by any NGS assembler and a set of long reads. The long reads are used to extend the contigs present in the NGS draft and possibly join overlapping contigs. EAGLER supports both PacBio and Oxford Nanopore reads.
The tool should be compatible with most UNIX flavors and has been successfully tested on the following operating systems:
- Mac OS X 10.11.1
- Mac OS X 10.10.3
- Ubuntu 14.04 LTS
To run the scaffolder correctly, the executables for the programs listed below should be reachable from the PATH variable of your shell.
- g++ (4.9.0 or later)
- GNU Make
- Burrows-Wheeler Aligner (0.7.12 or later)
- GraphMap Aligner (optional)
- Doxygen (optional)
The scaffolder depends directly on 2 libraries: SeqAn and cpppoa. Both libraries will be automatically downloaded, configured and built by entering the installation commands in a terminal window.
To install EAGLER run the following commands from the folder where you want to install the tool:
git clone https://github.com/mculinovic/EAGLER.git
cd EAGLER/
git submodule update --init --recursive
make
Running the make command without arguments will build the release version of the tool as the binary file release/eagler.
To build the debug version of the tool use:
make debug
To build both the debug and release versions use:
make all
Once the release version has been build you may run the following command to install the tool to /usr/local/bin:
make install
To delete all files generated during the build process, both for debug and release, use:
make clean
To remove the installed executable use:
make uninstall
The documentation for this tool was written to work with the doxygen documentation generator. To successfully generate the documentation, the doxygen executable must be reachable from your PATH variable.
To create the documentation in HTML and LaTeX format run the following command:
make docs
The HTML documentation is placed in docs/html, while the LaTeX documentation is placed in docs/latex. To view the HTML documentation open docs/html/index.html in any web browser. The PDF documentation is obtainable by compiling the generated LaTeX code with the provided makefile.
Use the following commands from the root of the project to create the PDF version of the documentation:
cd docs/latex/
make
open refman.pdf
Please check the links file for links to datasets and additional information on the alignment toolbox used by the scaffolder.
To run the tool please use the command format shown below:
./release/eagler [options] <draft_genome.fasta> <long_reads.fasta> <output_prefix/output_dir>
The implementation will automatically detect the number of hardware threads supported by the system.
The default configuration, i.e. no command line options, expects PacBio reads as input. The underlying aligner will be set to BWA and the Local/Global Realign algorithm will be used to extend the contigs.
To get a detailed view of the available options please run:
./release/eagler -h
###Arguments:
- draft_genome.fasta: FASTA file containing the draft genome created by some NGS pipeline
- long_reads.fasta: FASTA file containing long reads to be used in the scaffolding
- output_prefix/output_dir: the prefix to be added to the output files or the directory where the scaffolder should store the results
1) ./release/eagler -x pacbio -t 16 draft.fasta reads.fasta output_dir/
The above command will run the scaffolder over the draft genome draft.fasta using 24 parallel threads. The input for this example is a set of PacBio long reads from the reads.fasta file. The output of the scaffolder will consist of 3 files stored in the output_dir directory:
| Output File | Content |
|---|---|
| output_dir/contigs.fasta | Contigs from the draft genome extended by the scaffolder |
| output_dir/extensions.fasta | Left and right extensions for each contig in the draft |
| output_dir/scaffolds.fasta | Final scaffolds created by merging overlapping extended contigs |
2) ./release/eagler -g -x ont draft.fasta ont_reads.fasta example_2
The above command will run the scaffolder over the draft genome draft.fasta using as many parallel threads as there are cores on the host machine. In this case the input is a set of Oxford Nanopore 2D reads stored in the ont_reads.fasta file and the GraphMap aligner will be used to map them on the draft genome. The output of the scaffolder will consist of 3 files stored in the current working directory:
| Output File | Content |
|---|---|
| example_2.contigs.fasta | Contigs from the draft genome extended by the scaffolder |
| example_2.extensions.fasta | Left and right extensions for each contig in the draft |
| example_2.scaffolds.fasta | Final scaffolds created by merging overlapping extended contigs |
Some utility scripts are available in the scripts folder. All scripts have been developed and tested with Python 3.4.3.
For detailed usage instructions run the following command for the desired script:
python3 scripts/<script_name>.py --help
- genome2contigs: cuts a reference genome into multiple contigs
- reverse_complement: performs the reverse complement operation over sequences in a FASTA file
- extension_analysis: runs various statistics on contig extensions produced by the scaffolder