mkatari-bioinformatics-august-2013-gatknotes
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mkatari-bioinformatics-august-2013-gatknotes [2014/06/11 12:49] – created mkatari | mkatari-bioinformatics-august-2013-gatknotes [2014/06/13 07:23] – mkatari | ||
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bowtie2-build PTC_Human.fasta PTC_Human | bowtie2-build PTC_Human.fasta PTC_Human | ||
samtools faidx PTC_Human.fasta | samtools faidx PTC_Human.fasta | ||
- | java -jar / | + | java -jar / |
+ | R=PTC_Human.fasta | ||
+ | O=PTC_Human.dict | ||
</ | </ | ||
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* create a bam index for dedup bam files | * create a bam index for dedup bam files | ||
* realign samples | * realign samples | ||
+ | |||
+ | Once they are all processed | ||
* merge all samples to one bam file | * merge all samples to one bam file | ||
* sort and index this merged bam file | * sort and index this merged bam file | ||
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bowtie2 -x PTC_Human -U Cohen.fastq -S Cohen.sam | bowtie2 -x PTC_Human -U Cohen.fastq -S Cohen.sam | ||
samtools view -bS Cohen.sam > Cohen.bam | samtools view -bS Cohen.sam > Cohen.bam | ||
- | bowtie2 -x PTC_Human -U Sherman.fastq -S Sherman.sam | ||
- | samtools view -bS Sherman.sam > Sherman.bam | ||
</ | </ | ||
The picard method to sort is preferred by GATK | The picard method to sort is preferred by GATK | ||
< | < | ||
- | java -jar / | + | java -jar / |
- | java -jar / | + | INPUT=Cohen.bam |
+ | OUTPUT=Cohen.sorted.bam | ||
+ | SORT_ORDER=coordinate | ||
+ | |||
</ | </ | ||
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< | < | ||
- | java -jar / | ||
- | java -jar / | + | java -jar / |
+ | INPUT=Cohen.sorted.bam | ||
+ | OUTPUT=CohenRG.bam | ||
+ | RGLB=Cohen | ||
+ | RGPL=IonTorrent | ||
+ | RGPU=None | ||
+ | RGSM=Cohen | ||
</ | </ | ||
This will remove any reads that map to the same exact place. It is helpful to get rid of artifacts. | This will remove any reads that map to the same exact place. It is helpful to get rid of artifacts. | ||
< | < | ||
- | java -jar / | + | java -jar / |
+ | INPUT=CohenRG.bam | ||
+ | OUTPUT=Cohen.dedup.bam | ||
+ | METRICS_FILE=Cohen.dedup.metrics | ||
+ | REMOVE_DUPLICATES=TRUE | ||
+ | ASSUME_SORTED=TRUE | ||
- | java -jar / | ||
</ | </ | ||
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< | < | ||
samtools index Cohen.dedup.bam | samtools index Cohen.dedup.bam | ||
- | samtools index Sherman.dedup.bam | ||
# | # | ||
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-o CohenforIndelRealigner.intervals | -o CohenforIndelRealigner.intervals | ||
- | # | ||
- | java -Xmx2g -jar / | ||
- | -T RealignerTargetCreator \ | ||
- | -R PTC_Human.fasta \ | ||
- | -I Sherman.dedup.bam \ | ||
- | -o ShermanforIndelRealigner.intervals | ||
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-targetIntervals CohenforIndelRealigner.intervals \ | -targetIntervals CohenforIndelRealigner.intervals \ | ||
-o Cohen.dedup.realign.bam | -o Cohen.dedup.realign.bam | ||
- | |||
- | java -Xmx4g -jar / | ||
- | -T IndelRealigner \ | ||
- | -R PTC_Human.fasta \ | ||
- | -I Sherman.dedup.bam \ | ||
- | -targetIntervals ShermanforIndelRealigner.intervals \ | ||
- | -o Sherman.dedup.realign.bam | ||
</ | </ | ||
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< | < | ||
- | java -jar / | + | java -jar / |
+ | INPUT=Sherman.dedup.realign.bam | ||
+ | INPUT=Cohen.dedup.realign.bam | ||
+ | OUTPUT=ShermanCohenMerged.bam | ||
samtools sort ShermanCohenMerged.bam ShermanCohenMerged.sorted | samtools sort ShermanCohenMerged.bam ShermanCohenMerged.sorted | ||
+ | |||
samtools index ShermanCohenMerged.sorted.bam | samtools index ShermanCohenMerged.sorted.bam | ||
</ | </ | ||
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-glm SNP \ | -glm SNP \ | ||
-o PTC_human.gatk.vcf | -o PTC_human.gatk.vcf | ||
+ | |||
+ | </ | ||
+ | |||
+ | If you want to load the vcf file into IGV, remember to index it first. | ||
+ | < | ||
+ | module load igvtools | ||
+ | igvtools index PTC_human.gatk.vcf | ||
+ | </ | ||
+ | |||
+ | If you would like to generate a table of from the vcf file use the following command | ||
+ | < | ||
+ | java -jar GenomeAnalysisTK.jar \ | ||
+ | -R PTC_Human.fasta | ||
+ | -T VariantsToTable \ | ||
+ | -V PTC_human.gatk.vcf \ | ||
+ | -F CHROM -F POS -F ID -F QUAL -F AC \ | ||
+ | -GF GT -GF GQ \ | ||
+ | -o PTC_human.gatk.vcf.table | ||
</ | </ |
mkatari-bioinformatics-august-2013-gatknotes.txt · Last modified: 2016/08/17 08:37 by mkatari