GATK - k821209/pipelines GitHub Wiki

required

http://broadinstitute.github.io/picard/
GATK # https://www.amazon.com/clouddrive/folder/TMkWwhkPSi6fsuk2wI8l4Q
samtools
sambamba (for fast sort)

apt-get install littler # for Rscript

add read group

java -jar /program/picard-tools-2.1.1/picard.jar AddOrReplaceReadGroups INPUT=Pepper_1.55_R_reference_genome_R.taeahn.sorted.bam OUTPUT=Pepper_1.55_R_reference_genome_R.taeahn.sorted.bam.rg.bam LB=taeahn PL=Illumina PU=NA SM=taeahn VALIDATION_STRINGENCY=LENIENT

full pipe

https://www.biostars.org/p/8237/

#Create sequence dictionary
java -jar~/bin/picard-tools-1.8.5/CreateSequenceDictionary.jar REFERENCE=reference.fasta OUTPUT=reference.dict

#Align reads and assign read group
bwa mem -R “@RG\tID:FLOWCELL1.LANE1\tPL:ILLUMINA\tLB:test\tSM:PA01” reference.fasta R1.fastq.gz R2.fastq.gz > aln.sam

#Sort sam file
java -jar ~/bin/picard-tools-1.8.5/SortSam.jar I=aln.sam O=sorted.bam SORT_ORDER=coordinate

#Mark duplicates
java -jar ~/bin/picard-tools-version/MarkDuplicates.jar I=sorted.bam O=dedup.bam METRICS_FILE=metrics.txt

#Sort bam file
java -jar ~/bin/picard-tools-version/BuildBamIndex.jar INPUT=dedup.bam

#Create realignment targets
java -jar ~/bin/GATK3.3/GenomeAnalysisTK.jar -T RealignerTargetCreator -R reference.fasta -I dedup.bam -o targetintervals.list

#Indel realignment
java -jar ~/bin/GATK3.3/GenomeAnalysisTK.jar -T IndelRealigner -R PA01.fasta -I dedup.bam -targetIntervals targetintervals.list -o realigned.bam

#Call variants (HaplotypeCaller)
java -jar ~/bin/GATK3.3/GenomeAnalysisTK.jar -T HaplotypeCaller -R reference.fasta -I realigned.bam -ploidy 1 -stand_call_conf 30 -stand_emit_conf 10 -o raw.vcf

The resulting vcf file will contain your variant calls!

Then you can optionally filter the variants:

#Filter variants
~/bin/vcflib/bin/vcffilter -f ‘DP > 9’ -f ‘QUAL > 10’ raw.vcf > filtered.vcf

Or first split the raw.vcf file into SNPs and indels:

#Extract SNPs
java -jar ~/bin/GATK3.3/GenomeAnalysisTK.jar -T SelectVariants -R reference.fasta -V raw.vcf -selectType SNP -o snps.vcf

#Extract Indels
java -jar ~/bin/GATK/GenomeAnalysisTK.jar -T SelectVariants -R reference.fasta -V raw.vcf -selectType INDEL -o indels.vcf