ftarc

FASTQ-to-analysis-ready-CRAM Workflow Executor for Human Genome Sequencing

Installation

$ pip install -U ftarc

Dependent commands:

• pigz
• pbzip2
• bgzip
• tabix
• samtools (and plot-bamstats)
• gnuplot
• java
• gatk
• cutadapt
• fastqc
• trim_galore
• bwa or bwa-mem2

Docker image

Pull the image from Docker Hub.

$ docker image pull dceoy/ftarc

Usage

Create analysis-ready CRAM files from FASTQ files

input files	output files
read1/read2 FASTQ (Illumina)	analysis-ready CRAM

1. Download hg38 resource data.

   $ ftarc download --dest-dir=/path/to/download/dir

2. Write input file paths and configurations into ftarc.yml.

   $ ftarc init
   $ vi ftarc.yml  # => edit

   Example of ftarc.yml:

   ---
   reference_name: hs38DH
   adapter_removal: true
   metrics_collectors:
     fastqc: true
     picard: true
     samtools: true
   resources:
     reference_fa: /path/to/GRCh38_full_analysis_set_plus_decoy_hla.fa
     known_sites_vcf:
       - /path/to/Homo_sapiens_assembly38.dbsnp138.vcf.gz
       - /path/to/Mills_and_1000G_gold_standard.indels.hg38.vcf.gz
       - /path/to/Homo_sapiens_assembly38.known_indels.vcf.gz
   runs:
     - fq:
         - /path/to/sample01.WGS.R1.fq.gz
         - /path/to/sample01.WGS.R2.fq.gz
     - fq:
         - /path/to/sample02.WGS.R1.fq.gz
         - /path/to/sample02.WGS.R2.fq.gz
     - fq:
         - /path/to/sample03.WGS.R1.fq.gz
         - /path/to/sample03.WGS.R2.fq.gz
       read_group:
         ID: FLOWCELL-1
         PU: UNIT-1
         SM: sample03
         PL: ILLUMINA
         LB: LIBRARY-1

3. Create analysis-ready CRAM files from FASTQ files

   $ ftarc pipeline --yml=ftarc.yml --workers=2

   Standard workflow:

   1. Trim adapters
      • trim_galore
   2. Map reads to a human reference genome
      • bwa mem (or bwa-mem2 mem)
   3. Mark duplicates
      • gatk MarkDuplicates
      • gatk SetNmMdAndUqTags
   4. Apply BQSR (Base Quality Score Recalibration)
      • gatk BaseRecalibrator
      • gatk ApplyBQSR
   5. Remove duplicates
      • samtools view
   6. Validate output CRAM files
      • gatk ValidateSamFile
   7. Collect QC metrics
      • fastqc
      • samtools
      • gatk

Preprocessing and QC-check

• Validate BAM or CRAM files using Picard

  $ ftarc validate /path/to/genome.fa /path/to/aligned.cram

• Collect metrics from FASTQ files using FastQC

  $ ftarc fastqc read1.fq.gz read2.fq.gz

• Collect metrics from FASTQ files using FastQC

  $ ftarc samqc /path/to/genome.fa /path/to/aligned.cram

• Apply BQSR to BAM or CRAM files using GATK

  $ ftarc bqsr \
      --known-sites-vcf=/path/to/Homo_sapiens_assembly38.dbsnp138.vcf.gz \
      --known-sites-vcf=/path/to/Mills_and_1000G_gold_standard.indels.hg38.vcf.gz \
      --known-sites-vcf=/path/to/Homo_sapiens_assembly38.known_indels.vcf.gz \
      /path/to/genome.fa /path/to/markdup.cram

• Remove duplicates in marked BAM or CRAM files

  $ ftarc dedup /path/to/genome.fa /path/to/markdup.cram

Run ftarc --help for more information.