Getting Started

Introduction

The CARLISLE Pipeline begins with raw FASTQ files and performs trimming followed by alignment using BOWTIE2. Data is then normalized through either the use of an user-species species (IE E.Coli) spike-in control or through the determined library size. Peaks are then called using MACS2, SEACR, and GoPEAKS with various options selected by the user. Peaks are then annotated, and summarized into reports. If designated, differential analysis is performed using DESEQ2. QC reports are also generated with each project using FASTQC and MULTIQC. Annotations are added using HOMER and ROSE. GSEA Enrichment analysis predictions are added using CHIPENRICH.

The following are sub-commands used within CARLISLE:

  • initialize: create a working directory for the pipeline to run
  • dryrun: predict the binding of peptides to any MHC molecule
  • cluster: execute the pipeline on the Biowulf HPC
  • local: execute a local, interactive, session
  • git: execute GitHub actions
  • unlock: unlock directory
  • DAG: create DAG report
  • report: create SNAKEMAKE report
  • runtest: copies test manifests and files to WORKDIR

Setup Dependencies

CARLISLE has several dependencies listed below. These dependencies can be installed by a sysadmin. All dependencies will be automatically loaded if running from Biowulf.

  • bedtools: "bedtools/2.30.0"
  • bedops: "bedops/2.4.40"
  • bowtie2: "bowtie/2-2.4.2"
  • cutadapt: "cutadapt/1.18"
  • fastqc: "fastqc/0.11.9"
  • fastq_screen: "fastq_screen/0.15.2"
  • fastq_val: "/data/CCBR_Pipeliner/iCLIP/bin/fastQValidator"
  • fastxtoolkit: "fastxtoolkit/0.0.14"
  • gopeaks: "github clone https://github.com/maxsonBraunLab/gopeaks"
  • macs2: "macs/2.2.7.1"
  • multiqc: "multiqc/1.9"
  • perl: "perl/5.34.0"
  • picard: "picard/2.26.9"
  • python37: "python/3.7"
  • R: "R/4.2.2"
  • rose: "ROSE/1.3.1"
  • samtools: "samtools/1.15"
  • seacr: "seacr/1.4-beta.2"
  • ucsc: "ucsc/407"

Login to the cluster

CARLISLE has been exclusively tested on Biowulf HPC. Login to the cluster's head node and move to your project directory.

# ssh into cluster's head node
ssh -Y $USER@biowulf.nih.gov

Load an interactive session

An interactive session should be started before performing any of the pipeline sub-commands, even if the pipeline is to be executed on the cluster.

# Grab an interactive node
sinteractive --time=12:00:00 --mem=8gb  --cpus-per-task=4 --pty bash

Load dependencies

Load the ccbrpipeliner module to access CARLISLE:

module load ccbrpipeliner

or specify an exact version of the module:

module load ccbrpipeliner/7

CARLISLE will then be in your path and you can check which version you have loaded:

carlisle --version