Informatics for High-throughput Sequencing Data Analysis 2019 Integrated Assignment

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CBW HT-seq Integrative Assignment

Written originally by Mathieu Bourgey, edited by Florence Cavalli

Task

We will perform the same analysis as in Module 3 but using the mother and father samples i.e sample NA12891 and NA12891.

The fastq files are in the following directory of the cloud instance: ~/CourseData/HT_data/Module3/

 * raw_reads/NA12891/NA12891_CBW_chr1_R1.fastq.gz
 * raw_reads/NA12891/NA12891_CBW_chr1_R2.fastq.gz
 * raw_reads/NA12892/NA12892_CBW_chr1_R1.fastq.gz
 * raw_reads/NA12892/NA12892_CBW_chr1_R2.fastq.gz

Environment setup

# Start a fresh docker container:

docker run --privileged -v /tmp:/tmp --network host -it -w $PWD -v $HOME:$HOME -v /media:/media --user $UID:$GROUPS -v /etc/group:/etc/group -v /etc/passwd:/etc/passwd c3genomics/genpipes:0.8

Then inside the docker container:

#set up environment variables
export REF=$WORK_DIR/reference/
export WORK_DIR=~/workspace/HTseq/Integrative_Assignment/

rm -rf $WORK_DIR
mkdir -p $WORK_DIR
cd $WORK_DIR
ln -fs ~/CourseData/HT_data/Module3/* .

# Load the software modules

module load mugqic/java/openjdk-jdk1.8.0_72 mugqic/bvatools/1.6 mugqic/trimmomatic/0.36 mugqic/samtools/1.9 mugqic/bwa/0.7.17 mugqic/GenomeAnalysisTK/4.1.0.0 mugqic/R_Bioconductor/3.5.0_3.7

Task list:

  1. Check read QC

  2. Trim unreliable bases from the read ends

  3. Align the reads to the reference

  4. Sort the alignments by chromosome position

  5. Realign short indels

  6. Fixe mate issues (optional)

  7. Mark duplicates

  8. Recalibrate the Base Quality

  9. Generate alignment metrics

Discussion/Questions:

  1. Explain the purpose of each step

  2. Which software tool can be used for each step

The full commands can be downloaded here solution