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NanoSwe: Analysing PromethION Sequencing Data of Swedish Genomes 🇸🇪

Introduction

NanoSwe is a preliminary analysis toolkit for experiments that involve sequencing data from ONT's PromethION device. It has also been used for other long-read SweGen data (e.g. PacBio).

Bioinformatics ToolKit

Purpose Program
Quality Control NanoPlot for QC and NanoComp
Mapping to the reference Minimap2-2.14
Sorting, Indexing, and calculating statistics Samtools 1.9
Subsampling Sambamba 0.7.1
BAM QC Statistics Qualimap 2.2.1
Structural Variant Calling Sniffles 1.0.10
Data Extraction (VCF Files only) bcftools 1.9
Finding intersection in genomic regions Survivor 1.0.7
Evaluation of SVs Survivor 1.0.7 and surpyvor: 0.5.0
Removing control DNA sequences NanoLyse
Trimming Short Reads BBMap/BBTools
Homology Detection Blast 2.7.1+
Data Visualisation R version 3.5.3. See the scripts directory for information on libraries/packages used.

Nanopore Data: Post-Sequencing

Example tree structure of nanopore sequencing data files

├── /basecalled/<sample>/<flowcell>/
│   ├── fastq_0.fastq
│   ├── fastq_850.fastq
│   ├── sequencing_summary_0.txt
│   ├── sequencing_summary_850.txt
│   └── reads (1)
│       ├── 0 (2)
│       │   ├── file_read_1_ch_90_strand.fast5
│       │   ├── file_read_41_ch_40_strand2.fast5
│       │   └── file_read_300_ch_40_strand2.fast5
│       └── 850
│           ├── file_read_1000_ch_200_strand.fast5
│           ├── file_read_9000_ch_100_strand.fast5
│           └── file_read_95000_ch_1000_strand2.fast5
└── /bin/

(1) Each folder contains ~8000 fast5 files
(2) fast5 file named e.g. PCT0001_YYYYMMDD_0001A20B002222C_{flowcell}_sequencing_run_{library_full_name}__read_{number}_ch_{number}_strand.fast5)

Nanopore Data: Post-Tidying

Example tree structure of data organisation

├── /basecalled/<sample>/<flowcell>/
│   ├── FASTQ_files
│   │   ├── fastq_0.fastq
│   │   └── fastq_850.fastq
│   ├── sequencing_summary
│   │   ├── sequencing_summary_0.txt
│   │   └── sequencing_summary_850.txt
│   ├── reads *
│   │   ├── 0 *
│   │   │   ├── file_read_1_ch_90_strand.fast5
│   │   │   ├── file_read_41_ch_40_strand2.fast5
│   │   │   └── file_read_300_ch_40_strand2.fast5
│   │   └── 850
│   │       ├── file_read_1000_ch_200_strand.fast5
│   │       ├── file_read_9000_ch_100_strand.fast5
│   │       └── file_read_95000_ch_1000_strand2.fast5
│   └── <sample>_analysis
│       ├── reference_genome.fna
|       ├── reference_genome.fna.fai
│       ├── Snakefile
│       ├── /bam_files/
│       ├── /vcf_files/
│       └── /logs/
└── /bin/

Sub-folder content

./scRipts - R scripts created for visulisation of long read data.
commands.md - Tool commands used for different analyses.

Data Sources

Recommended Readings

Recommended Pipeline(s)

Citation

If you plan to use repository as a guide, simply and kindly mention the link https://github.com/Nazeeefa/NanoSwe for acknowledgment. To cite our publication, you can cite it as as shown below otherwise visit citeas.org to choose a different format. Thank you.

AMA Style

Fatima N, Petri A, Gyllensten U, Feuk L, Ameur A. Evaluation of Single-Molecule Sequencing Technologies for Structural Variant Detection in Two Swedish Human Genomes. Genes. 2020; 11(12):1444.

Chicago Style

Fatima, Nazeefa; Petri, Anna; Gyllensten, Ulf; Feuk, Lars; Ameur, Adam. 2020. "Evaluation of Single-Molecule Sequencing Technologies for Structural Variant Detection in Two Swedish Human Genomes." Genes 11, no. 12: 1444.