Oxford Nanopore Minion Quality Control

Author

Michael Hall

Date

06/28/2022

Arabidopsis Thalania

Standard Workflow

MinionQC

Clone the Github Repository

git clone https://github.com/PBGLMichaelHall/MinionQC.git

Change Working Directory

cd/ MinionQC

Create a new enviroment

mamba env create –file env/MinionQCenv.yaml

Activate new Environment

conda activate MinionQC

Run Minion R Script

Rscript Minion_R_QC_Script/MinIONQC.R -i summary/sequencing_summary.txt -q 0

Data Tree Structure

(base) michael@mh-ubuntu:~/Desktop/Arabidopsis_Thalania_Run00123$ tree
.
├── fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_0.fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_10.fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_11.fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_12.fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_13.fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_14.fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_15.fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_16.fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_17.fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_18.fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_19.fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_1.fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_20.fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_21.fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_22.fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_23.fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_24.fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_25.fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_2.fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_3.fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_4.fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_5.fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_6.fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_7.fast5
│   ├── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_8.fast5
│   └── AJI658_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_9.fast5
└── guppy_out
├── fail
│   ├── fastq_runid_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_0_0.fastq
│   ├── fastq_runid_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_1_0.fastq
│   ├── fastq_runid_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_2_0.fastq
│   ├── fastq_runid_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_3_0.fastq
│   ├── fastq_runid_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_4_0.fastq
│   ├── fastq_runid_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_5_0.fastq
│   └── fastq_runid_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_6_0.fastq
├── guppy_basecaller-core-dump-db
│   ├── attachments
│   ├── completed
│   ├── new
│   ├── pending
│   └── settings.dat
├── guppy_basecaller_log-2022-05-23_12-49-31.log
├── guppy_basecaller_log-2022-05-23_18-21-02.log
├── guppy_out
│   ├── guppy_basecaller-core-dump-db
│   │   ├── attachments
│   │   ├── completed
│   │   ├── new
│   │   ├── pending
│   │   └── settings.dat
│   ├── guppy_basecaller_log-2022-05-23_12-49-21.log
│   └── sequencing_summary.txt
├── pass
│   ├── fastq_runid_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_0_0.fastq
│   ├── fastq_runid_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_1_0.fastq
│   ├── fastq_runid_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_2_0.fastq
│   ├── fastq_runid_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_3_0.fastq
│   ├── fastq_runid_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_4_0.fastq
│   ├── fastq_runid_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_5_0.fastq
│   └── fastq_runid_535642224ed16dcaf6e8cab4fc0ebb5f60ddaf03_6_0.fastq
├── sequencing_telemetry.js
└── summary
    ├── MinIONQC.R
     └── sequencing_summary.txt

16 directories, 49 files

Guppy Basecaller Log Standard Output

2022-05-23 12:49:31.950838 [guppy/message] ONT Guppy basecalling software version 5.0.17+99baa5b27
config file:        /opt/ont/guppy/data/dna_r9.4.1_450bps_hac.cfg
model file:         /opt/ont/guppy/data/template_r9.4.1_450bps_hac.jsn
input path:         fast5/
save path:          guppy_out/
chunk size:         2000
chunks per runner:  256
minimum qscore:     9
records per file:   4000
fastq compression:  ON
num basecallers:    1
cpu mode:           ON
threads per caller: 4

Conda/Mamba Environment Preview

name: MinionQC

channels:
  - default
  - bioconda
  - conda-forge
  - r

dependencies:
  - r-data.table
  - r-futile.logger
  - r-ggplot2
  - r-optparse
  - r-plyr
  - r-readr
  - r-reshape2
  - r-scales
  - r-viridis
  - r-yaml

Standard R Script Output

mamba env create --file MinionQCenv.yaml

conda activate MinionQC

Rscript MinIONQC.R -i sequencing_summary.txt -q 0


INFO [2022-06-28 10:59:29] Loading input file: sequencing_summary.txt
INFO [2022-06-28 10:59:30] MinION flowcell detected
INFO [2022-06-28 10:59:30] .: creating output directory:.
INFO [2022-06-28 10:59:30] .: summarising input file for flowcell
INFO [2022-06-28 10:59:30] .: plotting length histogram
INFO [2022-06-28 10:59:32] .: plotting mean Q score histogram
INFO [2022-06-28 10:59:33] .: plotting flowcell overview
INFO [2022-06-28 10:59:39] .: plotting flowcell yield over time
INFO [2022-06-28 10:59:40] .: plotting flowcell yield by read length
INFO [2022-06-28 10:59:41] .: plotting sequence length over time
INFO [2022-06-28 10:59:44] .: plotting Q score over time
INFO [2022-06-28 10:59:46] .: plotting reads per hour
INFO [2022-06-28 10:59:47] .: plotting read length vs. q score scatterplot
INFO [2022-06-28 10:59:49] .: plotting flowcell channels summary histograms
INFO [2022-06-28 10:59:50] .: plotting physical overview of output per channel

Figures/Plots