GotCloud: Variant Calling Pipeline

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The Variant Calling Pipeline (UMAKE) takes recalibrated BAM files and detects SNPs and calls their genotypes, producing VCF files.

Input Data:

• Aligned/Processed/Recalibrated BAM files
• Index file containing Sample IDs & BAM file names
• Reference files
• (Optional) Configuration file to override default options

Configuration File

A default configuration file is automatically loaded. Users must specify their own configuration file specifying just the values different than the defaults.

Comments begin with a #

Format: KEY = value

Where KEY is the item being set and value is its new value

Required User Config Files Settings

The following Config File Settings must be specified by the user:

• CHRS = # space separated list of chromosomes you want
• BAM_INDEX = # path to the Index File of BAMs

Required on Command-Line or in Config File

The following Command-Line or Config File Settings must be specified by the user:

• --outdir/OUTDIR= # path to desired output directory

Targeted/Exome Sequencing Settings

If you are running Targeted/Exome Sequencing, the user should specify:

• Write loci file when performing pileup
  • WRITE_TARGET_LOCI = TRUE
• Specify the directory to store target information, for example: targetDir
  • TARGET_DIR = targetDir

If all individuals have the same target:

• Specify the single bed file, for example: target.bed
  • UNIFORM_TARGET_BED = target.bed

If not all individuals have the same target:

• Specify the file containing the sample id -> bed map, for example: targetMap.txt
  • MULTIPLE_TARGET_MAP = targetMap.txt
    • Each line of the file contains [SM_ID] [TARGET_BED]

Optional Settings:

• Extend the target region by a given number of bases, for example: 50
  • OFFSET_OFF_TARGET = 50
• Exclude off-target regions when using samtools view (may make command line too long)
  • SAMTOOLS_VIEW_TARGET_ONLY = TRUE

Reference Files

• Reference Sequence in fasta format.
  • REF = path/file.fa
• Indel VCF File Prefix
  • INDEL_PREFIX = path/indels.sites.hg19
  • path/ contains indels.sites.hg19.chr20.vcf for each chromosome being processed
• DBSNP File Prefix
  • DBSNP_PREFIX = path/dbsnp_135_b37.rod
  • path/ contains dbsnp_135_b37.rod.chr20.map for each chromosome being processed
• HapMap3 polymorphic site prefix
  • HM3_PREFIX = path/hapmap3.qc.poly
  • path/ contains hapmap3.qc.poly.chr20.bim & hapmap3.qc.poly.chr20.frq for each chromosome being processed

Can be downloaded from: [| FTP Download of Full Resource Files]

INDEL_PREFIX = $(UMAKE_ROOT)/ref/indels/1kg.pilot_release.merged.indels.sites.hg19 # 1000 Genomes Pilot 1 indel VCF prefix DBSNP_PREFIX = $(UMAKE_ROOT)/ref/dbSNP/dbsnp_135_b37.rod # dbSNP file prefix HM3_PREFIX = $(UMAKE_ROOT)/ref/HapMap3/hapmap3_r3_b37_fwd.consensus.qc.poly # HapMap3 polymorphic site prefix

Chromosome X Calling

• PED_INDEX = pedfile.ped

Running

Running umake is straightforward:

cd ~/myseq
/usr/local/biopipe/bin/umake --conf myconf ???
make -f [out-prefix].Makefile -j [# parallel jobs]