NanoQC

NanoQC is a bioinformatics pipeline that performs quality control of basecalled Nanopore sequence data in .fastq.gz format.

Overview

1. Perform read QC with FastQC.
2. Perform read QC with NanoPlot.
3. Remove adapters with Porechop_ABI.
4. Collect QC reports with MultiQC.

Usage

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.

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.

NanoQC can be run using two different input types:

• A samplesheet, including sample names and paths to Nanopore-basecalled gzipped FASTQ files, or
• A folder containing Nanopore-basecalled gzipped FASTQ files.

Input type: Samplesheet

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

samplesheet.csv:

sample,fastq
CONTROL_1,data/AEG588A1.fastq.gz
CONTROL_2,data/AEG588A2.fastq.gz
TREATMENT_1,data/AEG575A5.fastq.gz

Each row represents a gzipped FASTQ file.

Now, you can run the pipeline using:

nextflow run pcrxn/nanoqc \
   -profile <docker/singularity/.../institute> \
   --input samplesheet.csv \
   --outdir <OUTDIR>

Input type: Folder

Instead of a samplesheet, the user can instead provide a path to a directory containing gzipped FASTQ files. In this case, the sample name will be the name of the file up until the first period (.).

For example, for a folder data/ that looks as follows:

data
├── ERR9958133.fastq.gz
└── ERR9958134.fastq.gz

The pipeline can be run using:

nextflow run pcrxn/nanoqc \
   -profile <docker/singularity/.../institute> \
   --input_folder data/ \
   --outdir <OUTDIR>

If the names of the gzipped FASTQ files do not end with .fastq.gz, an alternate extension can be specified using --extension.

Contributions and support

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

Citations

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

This pipeline uses code and infrastructure developed and maintained by the nf-core initative, and reused here under the MIT license.

Ewels PA, Peltzer A, Fillinger S, Patel H, Alneberg J, Wilm A, Garcia MU, Di Tommaso P, Nahnsen S. The nf-core framework for community-curated bioinformatics pipelines. Nat Biotechnol. 2020 Mar;38(3):276-278. doi: 10.1038/s41587-020-0439-x. PubMed PMID: 32055031.

In addition, references of tools and data used in this pipeline are as follows:

• FastQC

  Andrews S. (2010). FastQC: A Quality Control Tool for High Throughput Sequence Data [Online].

• MultiQC

  Ewels P, Magnusson M, Lundin S, Käller M. MultiQC: summarize analysis results for multiple tools and samples in a single report. Bioinformatics. 2016 Oct 1;32(19):3047-8. doi: 10.1093/bioinformatics/btw354. Epub 2016 Jun 16. PubMed PMID: 27312411; PubMed Central PMCID: PMC5039924.

• NanoPlot

  De Coster W, Rademakers R. NanoPack2: population-scale evaluation of long-read sequencing data, Bioinformatics. 2023 May 12;39(5):btad311. doi: 10.1093/bioinformatics/btad311.

• Porechop_ABI

  Bonenfant Q, Noé L, Touzet H. Porechop_ABI: discovering unknown adapters in Oxford Nanopore Technology sequencing reads for downstream trimming, Bioinformatics Advances. 2022 Nov 21;3(1):vbac085. doi: 10.1093/bioadv/vbac085.