Introduction

nf-core/genomeassembler is a bioinformatics pipeline that carries out genome assembly, polishing and scaffolding from long reads (ONT or pacbio). Assembly can be done via flye or hifiasm, polishing can be carried out with medaka (ONT), or pilon (requires short-reads), and scaffolding can be done using LINKS, Longstitch, or RagTag (if a reference is available). Quality control includes BUSCO, QUAST and merqury (requires short-reads). Currently, this pipeline does not implement phasing of polyploid genomes or HiC scaffolding.

Usage

Note

If you are new to Nextflow and nf-core, please refer to this page on how to set-up Nextflow. Make sure to test your setup with -profile test before running the workflow on actual data.

First, prepare a samplesheet with your input data that looks as follows:

samplesheet.csv:

sample,ontreads,hifireads,ref_fasta,ref_gff,shortread_F,shortread_R,paired
sampleName,ontreads.fa.gz,hifireads.fa.gz,assembly.fasta.gz,reference.fasta,reference.gff,short_F1.fastq,short_F2.fastq,true

Each row represents one genome to be assembled. sample should contain the name of the sample, ontreads should contain a path to ONT reads (fastq.gz), hifireads a path to HiFi reads (fastq.gz), ref_fasta and ref_gff contain reference genome fasta and annotations. shortread_F and shortread_R contain paths to short-read data, paired indicates if short-reads are paired. Columns can be omitted if they contain no data, with the exception of shortread_R, which needs to be present if shortread_F is there, even if it is empty.

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Now, you can run the pipeline using:

nextflow run nf-core/genomeassembler \
   -profile <docker/singularity/.../institute> \
   --input samplesheet.csv \
   --outdir <OUTDIR>

Warning

Please provide pipeline parameters via the CLI or Nextflow -params-file option. Custom config files including those provided by the -c Nextflow option can be used to provide any configuration except for parameters; see docs.

For more details and further functionality, please refer to the usage documentation and the parameter documentation.

Pre-set profiles

To ease configuration, there are a couple of pre-defined profiles for various combinations of read sources and assemblers (named readtype_assembler)

ONT	HiFI	Assembly-strategy	Profile name
Yes	No	flye	ont_flye
No	Yes	flye	hifi_flye
No	Yes	hifiasm	hifi_hifiasm
Yes	Yes	hifiasm --ul	hifiont_hifiasm
Yes	Yes	Scaffolding of ONT assemblies onto HiFi assemblies	hifiont_flyehifiasm

Pipeline specific parameters

Parameter	Description	Type	Default	Required	Hidden
ont	ONT reads available?	boolean
hifi	HiFi reads available?	boolean
short_reads	Short reads available?	boolean
collect	collect ONT reads into a single file	boolean
porechop	run porechop on ONT reads	boolean
lima	run lima on HiFi reads?	boolean
pacbio_primers	file containing pacbio primers for trimming with lima	string
trim_short_reads	trim short reads with trimgalore	boolean
assembler	Assembler to use. Valid choices are: 'hifiasm', 'flye', or 'flye_on_hifiasm'. flye_on_hifiasm will scaffold flye assembly (ont) on hifiasm (hifi) assembly using ragtag	string
kmer_length	kmer length to be used for jellyfish	integer
read_length	read length for genomescope (ONT only)	string
dump	dump jellyfish output	boolean
meryl_k	kmer length for meryl	integer
use_ref	use reference genome	boolean
genome_size	expected genome size	string
flye_mode	flye mode	string	"--nano-hq"
flye_args	additional args for flye	string	""
qc_reads	Long reads that should be used for QC when both ONT and HiFi reads are provided. Options are 'ONT' or 'HIFI'	string	"ONT"
hifiasm_ont	Use hifi and ONT reads with hifiasm --ul	boolean
hifiasm_args	Extra arguments passed to hifiasm	string	""
polish_pilon	Polish assembly with pilon?	boolean
polish_medaka	Polish assembly with medaka (ONT only)	boolean
medaka_model	model to use with medaka	string	'r1041_e82_400bps_hac_v4.2.0'
scaffold_ragtag	Scaffold with ragtag (requires reference)?	boolean
scaffold_links	Scaffolding with links?	boolean
scaffold_longstitch	Scaffold with longstitch?	boolean
lift_annotations	Lift-over annotations (requires reference)?	boolean
busco	Run BUSCO?	boolean
busoc_db	Path to busco db	string	''
busco_lineage	Busco lineage to use	string	"brassicales_odb10"
quast	Run quast	boolean
skip_assembly	skip assembly steps Help Skip assembly and perform only qc.	boolean
skip_alignments	skip alignments during qc	boolean
jellyfish	run jellyfish and genomescope on ONT reads to compute k-mer distribution and estimate genome size	boolean
yak	run qc via yak	boolean

Pipeline output

To see the results of an example test run with a full size dataset refer to the results tab on the nf-core website pipeline page. For more details about the output files and reports, please refer to the output documentation.

Credits

nf-core/genomeassembler was originally written by Niklas Schandry (@nschan).

We thank the following people for their extensive assistance in the development of this pipeline:

• Mahesh Binzer-Panchal (@mahesh-panchal)
• Matthias Hörtenhuber (@mashehu)
• Daniel Straub (@d4straub)

Contributions and Support

If you would like to contribute to this pipeline, please see the contributing guidelines.

For further information or help, don't hesitate to get in touch on the Slack #genomeassembler channel (you can join with this invite).

Citations

An extensive list of references for the tools used by the pipeline can be found in the CITATIONS.md file.

You can cite the nf-core publication as follows:

The nf-core framework for community-curated bioinformatics pipelines.

Philip Ewels, Alexander Peltzer, Sven Fillinger, Harshil Patel, Johannes Alneberg, Andreas Wilm, Maxime Ulysse Garcia, Paolo Di Tommaso & Sven Nahnsen.

Nat Biotechnol. 2020 Feb 13. doi: 10.1038/s41587-020-0439-x.