Catch-up

Environment setup

Directories and symlinks

Directories and links needed in your home directory.

cd 
ln -s -f $SCRATCH scratch
ln -s -f $WORK2 work2
ln -s -f /work2/projects/BioITeam
ln -s -f /work2/projects/BioITeam/projects/courses/Core_NGS_Tools CoreNGS

mkdir -p ~/local/bin
cd ~/local/bin
ln -s -f /work2/projects/BioITeam/common/bin/launcher_creator.py
ln -s -f /work2/projects/BioITeam/common/bin/launcher_maker.py

.bashrc setup

If you already have a .bashrc set up, make a backup copy first. You can restore your original login script after class is over.

cd
cp .bashrc .bashrc.beforeNGSTools

Copy and configure the login profile for this class

cd
cp /work2/projects/BioITeam/projects/courses/Core_NGS_Tools/tacc/bashrc.corengs.stampede2  .bashrc
chmod 600 .bashrc

# or, using your symlink
cd
cp ~/CoreNGS/tacc/bashrc.corengs.stampede2  .bashrc
chmod 600 .bashrc

Source it to make it active (if this doesn't work, log off then log back in):

Copy a pre-configured login script

source ~/.bashrc

Environment variables

General

export ALLOCATION=UT-2015-05-18
export BIWORK=/work2/projects/BioITeam
export CORENGS=$BIWORK/projects/courses/Core_NGS_Tools

export PATH=.:$HOME/local/bin:$PATH

Turn on coloring by file type in the shell:

# For better colors using a dark background terminal, un-comment this line:
export LS_COLORS=$LS_COLORS:'di=1;33:fi=01:ln=01;36:'

# For better colors using a white background terminal:
export LS_COLORS=$LS_COLORS:'di=1;34:fi=01:ln=01;36:'

# May or may not be needed
export LS_OPTIONS='-N --color=auto -T 0

TACC intro

Commands files

Simple commands

mkdir -p $SCRATCH/core_ngs/slurm/simple
cd $SCRATCH/core_ngs/slurm/simple
cp $CORENGS/tacc/simple.cmds .

Wayness commands

mkdir -p $SCRATCH/core_ngs/slurm/wayness
cd $SCRATCH/core_ngs/slurm/wayness
cp $CORENGS/tacc/wayness.cmds .

Start an idev session

To start a 3-hour idev (interactive development) session:

Start an idev session

idev -p normal -m 120 -N 1 -n 68 -A UT-2015-05-18 --reservation=BIO_DATA_week_1

You can tell you're in a idev session because the hostname command will return a compute node name (e.g. c401-041.stampede2.tacc.utexas.edu) instead of a login node name (e.g. login2.stampede2.tacc.utexas.edu).

The n idev session will terminate when the requested time has expired, or you use the exit command.

Working with FASTQ

Yeast data

Working with some yeast ChIP-seq FASTQ data:

# Create a $SCRATCH area to work on data for this course,
# with a sub-direct[1ory for pre-processing raw fastq files
mkdir -p $SCRATCH/core_ngs/fastq_prep

# Make symbolic links to the original yeast data:
cd $SCRATCH/core_ngs/fastq_prep
ln -s -f $CORENGS/yeast_stuff/Sample_Yeast_L005_R1.cat.fastq.gz
ln -s -f $CORENGS/yeast_stuff/Sample_Yeast_L005_R2.cat.fastq.gz

# Copy over a small FASTQ file
cd $SCRATCH/core_ngs/fastq_prep
cp $CORENGS/misc/small.fq .

ATACseq data for MultiQC

Get some FastQC reports for MultiQC:

mkdir -p $SCRATCH/core_ngs/multiqc/fqc.atacseq
cd $SCRATCH/core_ngs/multiqc/fqc.atacseq
cp $CORENGS/multiqc/fqc.atacseq/*.html .

FASTQ files for cutadapt

For command-line cutadapt exploration:

cd $SCRATCH/core_ngs/fastq_prep
cp $CORENGS/human_stuff/Sample_H54_miRNA_L004_R1.cat.fastq.gz .
cp $CORENGS/human_stuff/Sample_H54_miRNA_L005_R1.cat.fastq.gz .
zcat Sample_H54_miRNA_L004_R1.cat.fastq.gz | head -2000 > miRNA_test.fq

For batch cutadapt processing:

mkdir -p $SCRATCH/core_ngs/cutadapt
cd $SCRATCH/core_ngs/cutadapt
cp $CORENGS/human_stuff/Sample_H54_miRNA_L004_R1.cat.fastq.gz .
cp $CORENGS/human_stuff/Sample_H54_miRNA_L005_R1.cat.fastq.gz .
cp $CORENGS/yeast_stuff/Yeast_RNAseq_L002_R1.fastq.gz .
cp $CORENGS/yeast_stuff/Yeast_RNAseq_L002_R2.fastq.gz .
cp $CORENGS/tacc/cuta.cmds .

Alignment workflow

Alignment workflow setup

Starting files:

# FASTA (for building references)
mkdir -p $SCRATCH/core_ngs/references/fasta
cp $CORENGS/references/*.* $SCRATCH/core_ngs/references/fasta/

# FASTQ (to align)
mkdir -p $SCRATCH/core_ngs/alignment/fastq
cp $CORENGS/alignment/*fastq.gz $SCRATCH/core_ngs/alignment/fastq/

References

Get a copy of all references we build in the exercises (including FASTA):

mkdir -p $SCRATCH/core_ngs/references
rsync -ptlvrP $CORENGS/references/ $SCRATCH/core_ngs/references/

BWA PE alignment of yeast data

To jump into aligning PE yeast data with BWA

# Pre-built references
mkdir -p $SCRATCH/core_ngs/references
rsync -avrP $CORENGS/references/ $SCRATCH/core_ngs/references/

# FASTQ (to align)
mkdir -p $SCRATCH/core_ngs/alignment/fastq
cp $CORENGS/alignment/*fastq.gz $SCRATCH/core_ngs/alignment/fastq/

# Alignment directory
mkdir -p $SCRATCH/core_ngs/alignment/yeast_bwa
cd $SCRATCH/core_ngs/alignment/yeast_bwa
ln -s -f ../fastq
ln -s -f ../../references/bwa/sacCer3

module load biocontainers  # takes a while
module load bwa
module load samtools

samtools manipulation of aligned yeast data

To jump into post-alignment manipulation of the yeast_pairedend.bam with samtools:

mkdir -p $SCRATCH/core_ngs/alignment/yeast_bwa
cd $SCRATCH/core_ngs/alignment/yeast_bwa
cp $CORENGS/catchup/yeast_bwa/yeast_pairedend.bam .

module load biocontainers  # takes a while
module load samtools

# If the sorted, indexed BAM is needed:
cp $CORENGS/catchup/yeast_bwa/yeast_pairedend.sort* .

SAMTools and BEDTools

Setup for samtools

Setup for samtools exercises

mkdir -p $SCRATCH/core_ngs/samtools
cd $SCRATCH/core_ngs/samtools
cp $CORENGS/catchup/for_samtools/* .

module load biocontainers  # takes a while
module load samtools

Core NGS Tools