NextFlow Install

Using Gitpod for a Preconfigured Nextflow Development Environment

Gitpod provides a cloud-based development environment that can be accessed through your web browser. This environment is ideal for running Nextflow and is preconfigured for your convenience.

Requirements:

• A GitHub, GitLab, or BitBucket account (provides 50 hours of usage per month)
• A web browser (Google Chrome or Firefox)

Accessing the Gitpod Environment

To access the Gitpod environment:

1. Prefix the Github repository URL with gitpod.io/#. For example:

   gitpod.io/#https://github.com/sateeshperi/nextflow_tutorial.git

2. Alternatively, click the button below to launch directly:

   

3. Install the Gitpod browser extension by following the instructions here. This will add a green Gitpod button to each Git repository for easy access. Once installed, navigate to the training repository (https://github.com/sateeshperi/nextflow_tutorial.git), and click the green button.

Sign in to your Github account and authorize Gitpod. If using BitBucket or GitLab, sign into Gitpod and change your integration on the following page: gitpod.io/integrations.

Once signed in, Gitpod will load (skip prebuild, if asked). Remember to save the URL of your Gitpod environment for future access.

Exploring the Gitpod IDE

The Gitpod environment comes with a sidebar for customizing your environment and performing basic tasks, a terminal for running programs, and a main window for viewing and editing files. The environment comes preinstalled with Nextflow, Conda, and Docker.

To save your files, choose your file of interest, then either use the sidebar to select “File/Save As…”, or use your keyboard shortcut to save as, then choose “Show local”. This will open an explorer to choose where to save your file on your local machine.

Setting Up Your Workspace

1. Create the varcal environment based on the yml file:

   mamba install multiqc
   

   mamba env update -n base -f environment.yml
   

2. Download the reference genome and raw reads:

   bash data/fetch_raw_data.sh
   

3. Trim raw reads using trimmomatic:

   bash data/trim.sh
   

Reopening a Gitpod Session

Running workspaces are automatically stopped after 30 minutes of inactivity. You can reopen the environment by going to gitpod.io/workspaces, finding your previous environment, clicking the three-dot button to the right, and selecting “Open”.

You can also reopen a previous environment by using the saved URL from your previous Gitpod environment. Environments are saved for up to two weeks.

Alternatively, you can start a new workspace by clicking the green Gitpod button or using the Gitpod URL: gitpod.io/#https://github.com/sateeshperi/nextflow_tutorial.git.

mkdir -p ~/bin/
cd ~/bin/
curl -s https://get.nextflow.io | bash
chmod +x nextflow
echo 'PATH=~/bin:$PATH' >> ~/.bashrc
which nextflow      ## should point back to ~/bin
nextflow -v         ## check install by invoking Nextflow, getting version

conda install nf-core
# or
pip install nf-core

channels:

* bioconda
* conda-forge
  dependencies:
* fastqc
* trimmomatic
* bwa
* samtools
* bcftools
* multiqc
* graphviz

mamba env create -n varcal -f environment.yml

cd ~/
mkdir -p nextflow_tutorial/data/ref_genome
cd nextflow_tutorial/data/ref_genome
curl -L -o ecoli_rel606.fasta.gz ftp://ftp.ncbi.nlm.nih.gov/genomes/all/GCA/000/017/985/GCA_000017985.1_ASM1798v1/GCA_000017985.1_ASM1798v1_genomic.fna.gz
gunzip ecoli_rel606.fasta.gz
ls

cd ~/
mkdir -p nextflow_tutorial/data/untrimmed_fastq/
cd nextflow_tutorial/data/untrimmed_fastq/

curl -O ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR258/004/SRR2589044/SRR2589044_1.fastq.gz
curl -O ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR258/004/SRR2589044/SRR2589044_2.fastq.gz
curl -O ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR258/003/SRR2584863/SRR2584863_1.fastq.gz
curl -O ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR258/003/SRR2584863/SRR2584863_2.fastq.gz
curl -O ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR258/006/SRR2584866/SRR2584866_1.fastq.gz
curl -O ftp://ftp.sra.ebi.ac.uk/vol1/fastq/SRR258/006/SRR2584866/SRR2584866_2.fastq.gz

  conda activate varcal

  cd nextflow_tutorial/data/untrimmed_fastq/

  for infile in *_1.fastq.gz
  do
     base=$(basename ${infile} _1.fastq.gz)
     trimmomatic PE ${infile} ${base}_2.fastq.gz \
                  ${base}_1.trim.fastq.gz ${base}_1un.trim.fastq.gz \
                  ${base}_2.trim.fastq.gz ${base}_2un.trim.fastq.gz \
                  SLIDINGWINDOW:4:20 MINLEN:25 ILLUMINACLIP:NexteraPE-PE.fa:2:40:15
  done

  mkdir ../trimmed_fastq
  mv *.trim* ../trimmed_fastq
  cd ../trimmed_fastq
  ls

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