DeSeq-Free

DeSeq-Free (Whole genome Deep Sequencing analysis of Cell Free tumor DNA ) is a Snakemake workflow, aimed to analyze WGS of circulating cell-free DNA (cfDNA) in the plasma of cancer patients together with their matched germline and tumour samples in a reproducible, automated, and partially contained manner. It is implemented such that alternative or similar analysis can be added or removed.

Contents

• Using the DeSeq-Free workflow
• Documentation
• Input files
• Output files
• Test Dataset

Using the DeSeq-Free workflow

We assume that you already have conda installed, otherwise you can easily install it:

To install conda: https://docs.conda.io/projects/conda/en/latest/user-guide/install/linux.html

• Input:

  metafile (can be .xlsx or .csv) with raw fastq.gz data that looks as follows:

  sample, lane, fq1, fq2, type
  
  Sample1, lane1, S1_L001_R1_001.fastq.gz, S1_L001_R2_001.fastq.gz, 0
  Sample1, lane2, S1_L002_R1_001.fastq.gz, S1_L002_R2_001.fastq.gz, 0
  

  Each row represents a single-end fastq file. Rows with the same sample identifier are considered technical replicates and will be automatically merged. type refers to sample type (0= buffy coat, 1= plasma, 2=tumor).

  • Reference genome
    

    Before starting, a user need to download reference genome.

    Download from NCBI, Ensembl, or any other autorities

    wget https://ftp.ensembl.org/pub/release-100/fasta/homo_sapiens/dna/Homo_sapiens.GRCh38.dna.toplevel.fa.gz
    

    • Index reference genome for bwa-mem2
      

      Prepare indexed genome for bwa-mem2 to boost mapping. Refer to the bwa-mem2 instruction.
      

      Example code:

      ./bwa-mem2 index <in.fasta>
      Where 
      <in.fasta> is the path to reference sequence fasta file and 
      

• Code:

  git clone https://github.com/mdelcorvo/DeSeq-Free.git
  cd DeSeq-Free && conda env create -f envs/workflow.yaml
  conda activate DeSeq-Free_workflow
  
  snakemake --use-conda \
  --config \
  input=inputfile.xlsx \
  output=output_directory \
  genome=genome.fasta
  

Output files

• Somatic variant analysis
• Variant allele frequency
• Annotation of somatic variants
• Somatic signatures
• Analysis of somatic CNAs
• Fragment size analysis