Michael G. Campana, 2023-2026
Smithsonian's National Zoo & Conservation Biology Institute
This Nextflow [1] pipeline automates the alignment of Illumina sequencing reads against a reference genome and mitogenome using BWA-MEM [2], SAMtools [3-4], and the Genome Analysis Toolkit [5]. It optionally performs PSMC analysis [6], read trimming using AdapterRemoval v2 [7] and circular mitogenome alignment using CircularMapper [8].
Please cite:
Prado, N.A., Armstrong, E.E., Brown, J.L., Goldenberg, S.Z., Leimgruber, P., Pearson, V.R., Maldonado, J.E., Campana, M.G. 2023. Genomic resources for Asian (Elephas maximus) and African savannah elephant (Loxodonta africana) conservation and health research. Journal of Heredity. 114(5): 529–538. DOI: 10.1093/jhered/esad034.
This software is licensed under the Smithsonian Institution terms of use.
After installing Nextflow and Conda, download the pipeline using:
nextflow pull campanam/Elephants
The nextflow.config file included with this repository contains a standard profile for running the pipeline locally. See the Nextflow documentation for assistance in generating a configuration profile for your computing system. The parameters you will need to provide to execute the pipeline are listed in the params block. These are:
outdir: Path to the output directory
refseq: Path to the genome reference sequence
csi: Use csi index (e.g. for chr > 512 Mb) (true or false)
circular_mtDNA: Perform secondary independent circular alignment of mitochondrial DNA
mtDNA: Path to the mitochondrial reference sequence
mtDNA_ID: Name of mtDNA sequence in mitochondrial reference sequence file
samples: Path to the CSV detailing sequencing libraries (See below)
reads: Path to the folder containing the FASTQ read pairs
read_trimming: Trim reads using AdapterRemoval v2 (true or false)
keep_trimmed_reads: Retain trimmed/merged reads in fastq format (true or false)
trimparams: Parameters for read-trimming. This pipeline expects paired-read input, so do not collapse reads.
min_uniq_mapped: Minimum number of unique mapped reads to retain an alignment file
java_options: String of options for Java executables.
markDuplicates: Choice of "picard", "samtools" or "sambamba" for duplicate marking
mapq: Minimum mapping quality (MapQ) to retain alignment. Set to 0 to bypass this filter
gatk: Perform genotyping using GATK (true or false)
email: Email to send completion status. Set to "NULL" for no email.
psmc: Run PSMC analysis (true or false)
psmc_opts: Parameter line for PSMC program
psmc_mpileup_opts: BCFtools mpileup filter options for PSMC
psmc_vcfutils_opts: vcfutils.pl filter options for PSMC
psmc_psmcfa_opts: fq2psmcfa options for PSMC
psmc_bootstrap: Number of PSMC bootstrap replicates
psmc_plot_opts: Options for psmc_plot.pl
The pipeline expects a headered CSV file listing samples and libraries with the following columns:
Sample,RG,Library,PL,Read1,Read2,Adapter1,Adapter2
Sample is the name of the sample. RG is the unique read-group identification for each set of sequence reads for an individual. Library is in the individual library identification (in case a library was sequenced more than once). PL is the platform identifier (ILLUMINA, SOLID, LS454, HELICOS or PACBIO). Read1 and Read2 give the forward and reverse read file names. Adapter1 and Adapter2 give the expected forward and reverse adapter sequences.
NB: The adapter columns can be omitted if no read-trimming will be performed.
Execute the pipeline using the following command:
nextflow run campanam/Elephants -r main -c <config_file.config> -profile standard
- Di Tommaso, P., Chatzou, M., Floden, E.W., Prieto Barja, P., Palumbo, E., Notredame, C. (2017) Nextflow enables reproducible computational workflows. Nat Biotechnol, 35, 316–319. DOI: 10.1038/nbt.3820.
- Li, H. (2013) Aligning sequence reads, clone sequences and assembly contigs with BWA-MEM. arXiv, 1303.3997v2.
- Li, H., Handsaker, B., Wysoker, A., Fennell, T., Ruan, J., Homer, N., Marth, G., Abecasis, G., Durbin, R., 1000 Genome Project Data Processing Subgroup (2009) The Sequence Alignment/Map format and SAMtools. Bioinformatics, 25, 2078-2079. DOI: 10.1093/bioinformatics/btp352.
- Danecek, P., Bonfield, J.K., Liddle, J., Marshall, J., Ohan, V., Pollard, M.O., Whitwham, A., Keane, T., McCarthy, S.A., Davies, R.M., Li, H. (2021) Twelves years of SAMtools and BCFtools. GigaScience, 10, giab008. DOI: 10.1093/gigascience/giab008.
- McKenna, A., Hanna, M., Banks, E., Sivachenko, A., Cibulskis, K., Kernytsky, A., Garimella, K., Altshuler, D., Gabriel, S., Daly, M., DePristo, M.A. (2010) The Genome Analysis Toolkit: a MapReduce framework for analyzing next-generation DNA sequencing data. Genome Res, 20, 1297-1303. DOI: 10.1101/gr.107524.110.
- Li, H., Durbin, R. (2011) Inference of human population history from individual whole-genome sequences. Nature, 475, 493-496. DOI 10.1038/nature10231.
- Schubert, M., Lindgreen, S., Orlando, L. (2016) AdapterRemoval v2: rapid adapter trimming, identification, and read merging. BMC Research Notes, 9, 88. DOI: 10.1186/s13104-016-1900-2.
- Peltzer, A., Jäger, G., Herbig, A., Seitz, A., Kniep, C., Krause, J., Nieselt, K. (2016) EAGER: efficient ancient genome reconstruction. Genome Biology, 17, 60. DOI: 10.1186/s13059-016-0918-z.
Image Credit: Skip Brown. 2018. Smithsonian's National Zoo & Conservation Biology Institute. Smithsonian Institution. https://www.si.edu/object/asian-elephant:nzp_NZP-20180628-596SB.