Pipeline consists of 3 chief scripts:
- bbmap_index.nf - Makes indexes for your references and gives them appropriate identificator
- main.nf - includes following submodules:
- preprocessing
- mapping
- classify_reads
- assembly
- binning
- additional.nf:
- binrefinement
- bins classification
All processes and their parameters can be found in subscripts/process.nf
All information about submodules - in according nf scripts. Each submodule can be launched on its own as separate script!
- GTDB-Tk requires ~84G of external data that needs to be downloaded and unarchived:
wget https://data.gtdb.ecogenomic.org/releases/latest/auxillary_files/gtdbtk_data.tar.gz # mirror: https://data.ace.uq.edu.au/public/gtdb/data/releases/latest/auxillary_files/gtdbtk_data.tar.gz
tar xvzf gtdbtk_data.tar.gz
- Link GTDB to the "gtdbtk_data" folder in pipeline:
ln -s /path/to/gtdbtk /path/to/pipeline/gtdbtk_data
- CheckM DB requires ~1.4G of external data that needs to be downloaded and unarchived:
wget https://data.ace.uq.edu.au/public/CheckM_databases/checkm_data_2015_01_16.tar.gz
tar xvzf checkm_data_2015_01_16.tar.gz
- Link CheckM database to the "checkm_data" folder in pipeline:
ln -s /path/to/checkm_data /path/to/pipeline/checkm_data
- Kraken requires ~16Gb of external data that needs to be downloaded and unarchived:
wget https://genome-idx.s3.amazonaws.com/kraken/k2_standard_16gb_20231009.tar.gz
tar xvzf k2_standard_16gb_20231009.tar.gz
- Link Kraken2 db to the "kraken2_data" folder in pipeline:
ln -s /path/to/kraken2_data /path/to/pipeline/kraken2_data
- Install MetaPhlAn in the pipeline directory:
mamba create -p /path/to/pipeline/metaphlan -c bioconda metaphlan
- It requires ~15Gb of external data that needs to be downloaded:
mamba run -p /path/to/pipeline/metaphlan metaphlan --install
- Install it in the pipeline directory:
mamba create -p /path/to/pipeline/mw-env -c ursky metawrap-mg=1.3.2
- Make corrections to metaWRAP bin_refinement.sh script:
sed -i 's/(( $SIZE > 50000)) &&//g' /path/to/pipeline/mw-env/bin/metawrap-modules/bin_refinement.sh
nextflow run bbmap_index.nf \
-with-report \
-with-conda \
--ref host.fna,human.fna
nextflow run main.nf \
-with-conda \
-with-report \
--ref host.fna,human.fna \
--ref_number 1,2 \
--reads 'pipeline/fq_check/*{1,2}.fq' \
--maxbin2 --concoct --metabat2 \ # specify at least one binner
--output pipeline_results
Optional parameters of main.nf with their default options:
- fastp options
- --compress_level 7
- --poly_g_min_len 5
- --poly_x_min_len 10
- --cut_mean_quality 27
- --n_base_limit 0
- --average_qual 25
- --length_required 80
- --working_thread_n 12
- kraken2, bracken options
- --classification_lvl "S"
- bbmap options
- --host_dir "host"
- --human_dir "human"
- --bbmap_memory false
- --bbmap_cpus 4
- metawrap binning options
- --concoct false
- --metabat2 false
- --maxbin2 false
- metawrap bin refinement options
- metawrap_completion 50
- metawrap_contamination 30