Tutorial: EMMAX GotCloud STOM: Lecture 5

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STOM 2014 Workshop - Practical Sessions 5

Lecture 5

The slides describing the notes below are available here (PDF)

Basic Setup

  • To see the files for the session 5(,6, and 8), type
ls /data/stom2014/session5/

If you see any errors, please let me know now!

  • For convenience, let’s set some variables
export S5=/data/stom2014/session5
  • Also, let's relocate the output directory from the yesterday's class
mv ~/out ~/out_session2
  • And create a new output directory
mkdir ~/out
  • And check the input files
ls $S5/examples/

Preparing Input Files

  • Index file - See the example index file already prepared for this project
cat $S5/examples/index/chr7.CFTR.fastq.index
  • Configuration File - See the example configuration file below.
cat $S5/examples/index/chr7.CFTR.align.conf

INDEX_FILE = index/chr7.CFTR.fastq.index
###################
# References
REF_DIR = chr7Ref
AS = NCBI37
REF = $(REF_DIR)/hs37d5.chr7.fa
DBSNP_VCF =  $(REF_DIR)/dbsnp_135.b37.chr7.CFTR.vcf.gz
HM3_VCF = $(REF_DIR)/hapmap_3.3.b37.sites.chr7.CFTR.vcf.gz

Default options should be mostly fine in many other cases. In this example, because it is not genome-wide calling, reference files are modified to be chr7-specific

Run GotCloud Alignment Pipeline

  • Using the prepared input files, align the FASTQ files using the gotcloud alignment pipeline
$S5/gotcloud/gotcloud align --conf $S5/examples/index/chr7.CFTR.align.conf  --outDir ~/out/align --baseprefix $S5/examples
  • Check if the output BAMs and QC metrics are produced
ls ~/out/align/bams
ls ~/out/align/QCFiles/

Understanding the Output Files

  • To see the content of BAM file in the format of SAM specification, try
 samtools view -h ~/out/align/bams/NA06984.recal.bam | less
  • Check the summary QC metrics
cat ~/out/align/QCFiles/NA06984.qplot.stats
  • Check whether the sample is contaminated from verifyBamID output
cat ~/out/align/QCFiles/NA06984.genoCheck.selfSM
cat ~/out/align/QCFiles/NA12878.genoCheck.selfSM