Changes between Version 2 and Version 3 of SOPs/variant_calling_GATK


Ignore:
Timestamp:
01/16/14 15:47:47 (11 years ago)
Author:
gbell
Comment:

--

Legend:

Unmodified
Added
Removed
Modified

  • SOPs/variant_calling_GATK

    v2 v3  
    1010export PATH=/usr/lib/jvm/java-7-openjdk-amd64/bin:$PATH
    1111\\ \\
    12 '''Index the reference genome.''' [Need to do just once.]
     12'''Index the reference genome.''' [Need to do just once, with [[http://samtools.sourceforge.net/samtools.shtml|samtools]].]
    1313  * samtools faidx /path/to/genome/genome.fa
    1414\\
    15 '''Create a genome dictionary.''' [Need to do just once.]
     15'''Create a genome dictionary.''' [Need to do just once, with Picard's [[http://picard.sourceforge.net/command-line-overview.shtml#CreateSequenceDictionary|CreateSequenceDictionary]].]
    1616  * java -jar /usr/local/share/picard-tools/CreateSequenceDictionary.jar R=/path/to/genome/genome.fa O=/path/to/genome/genome.dict
    1717\\
    18 '''Validate VCF file or known variants''' (with GATK's ValidateVariants)
     18'''Validate VCF file or known variants''' (with GATK's [[http://www.broadinstitute.org/gatk/gatkdocs/org_broadinstitute_sting_gatk_walkers_variantutils_ValidateVariants.html|ValidateVariants]])
    1919  * java -jar /usr/local/gatk/GenomeAnalysisTK.jar -T ValidateVariants -R /path/to/genome/genome.fa --variant:VCF SNPs_from_NCBI.sorted.vcf \\
    2020Respond to errors (by correcting or removing problematic variants), run command again, etc., until validation is successful. \\
    … …  
    2222
    2323\\
    24 '''Align reads to genome with bwa'''
     24'''Align reads to genome with [[http://bio-bwa.sourceforge.net/bwa.shtml|bwa]]'''
    2525  * bsub "bwa aln /path/to/genome/bwa/genome Reads_1.fq > Reads_1.sai"
    2626  * bsub "bwa samse /path/to/genome/bwa/genome Reads_1.sai  Reads_1.fq > Reads_1.bwa.sam"
    2727\\
    28 '''Convert SAM to BAM, sort, and index'''
     28'''Convert SAM to BAM, sort, and index''' with [[http://samtools.sourceforge.net/samtools.shtml|samtools]]
    2929  * bsub /nfs/BaRC_Public/BaRC_code/Perl/SAM_to_BAM_sort_index/SAM_to_BAM_sort_index.pl Reads_1.bwa.sam
    3030\\
    3131'''Mark duplicates''' (multiple identical reads mapped to the same location). \\
    32 Run Picard Tools' MarkDuplicates on each sample \\
     32Run Picard Tools' [[http://picard.sourceforge.net/command-line-overview.shtml#MarkDuplicates|MarkDuplicates]] on each sample \\
    3333May Need "VALIDATION_STRINGENCY=LENIENT" if you get  \\
    3434Exception in thread "main" net.sf.samtools.SAMFormatException: SAM validation error: ERROR: ... MAPQ should be 0 for unmapped read. \\
    … …  
    3636\\
    3737'''Add Read Group header information to each BAM file''' (or GATK won't let you continue) \\
    38 Run Picard Tools' [[http://picard.sourceforge.net/command-line-overview.shtml#AddOrReplaceReadGroups|AddOrReplaceReadGroups] on each sample. \\
     38Run Picard Tools' [[http://picard.sourceforge.net/command-line-overview.shtml#AddOrReplaceReadGroups|AddOrReplaceReadGroups]] on each sample. \\
    3939Specify RGSM (Read Group sample), RGLB (Read Group Library), RGPL (Read Group platform), and RGPU (Read Group platform unit [e.g. run barcode])
    4040  * bsub java -jar /usr/local/share/picard-tools/AddOrReplaceReadGroups.jar I=Reads_1.bwa.dedup.bam O=Reads_1.bwa.dedup.good.bam RGSM=My_sample RGLB=My_project RGPL=illumina RGPU=none VALIDATION_STRINGENCY=LENIENT
    4141\\
    42 '''Index BAM file(s)''' (optional; for IGV viewing)
     42'''Index BAM file(s)''' with [[http://samtools.sourceforge.net/samtools.shtml|samtools]] (optional; for IGV viewing)
    4343  * bsub samtools index Reads_1.bwa.dedup.good.bam
    4444\\