Editing Sequence Analysis Practice 2011/03/10

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Overview

The aim for today's practice is to perform variant calling from sequence alignment files

Steps

0. SETTING UP ENVIRONMENTAL VARIABLES 

setenv BIN /home/hyun/thu/bin
setenv IN /home/hyun/thu/input
setenv REF /home/hyun/thu/ref

setenv OUT ~/seq/thursday/output
mkdir --p ${OUT}

1. EXON-TARGETTED DATA : COMPUTING GENOTYPE LIKELHOOD FROM BAM FILES 

${BIN}/samtools-hybrid pileup -g -f ${REF}/human_g1k_v37_chr20.fa ${OUT}/NA12878.exon.sample.deduped.bam > ${OUT}/NA12878.exon.sample.glf

2. EXON-TARGETTED DATA : VIEW THE GENOTYPE LIKELIHOOD FORMAT 

${BIN}/samtools-hybrid glfview ${OUT}/NA12878.exon.sample.glf | less

TYPE 'q' to finish

3. EXON-TARGETTED DATA : SINGLE-SAMPLE GENOTYPE CALLING using GLFSINGLE 

${BIN}/glfSingle --maxDepth 10000 --minMapQuality 20 -p 0.9 -g ${OUT}/NA12878.exon.sample.glf -b ${OUT}/NA12878.exon.sample.vcf

4. EXON-TARGETTED DATA : VIEW THE VCF FILES AND COUNT # OF SNPS 

less ${OUT}/NA12878.exon.sample.vcf

grep -v ^# ${OUT}/NA12878.exon.sample.vcf | wc -l

5. DEEP-COVERAGE GENOME : COMPUTE THE GENOTYPE LIKELIHOOD 

${BIN}/samtools-hybrid pileup -g -f ${REF}/human_g1k_v37_chr20.fa ${IN}/NA12878.highcov.sample.bam > ${OUT}/NA12878.highcov.sample.glf

6. DEEP-COVERAGE GENOME : SINGLE-SAMPLE VARIANT CALLING 

${BIN}/glfSingle --maxDepth 10000  --minMapQuality 20 -p 0.9 -g ${OUT}/NA12878.highcov.sample.glf -b ${OUT}/NA12878.highcov.sample.vcf

7. DEEP-COVERAGE GENOME : VIEW THE VCF FILES AND COUNT # OF SNPS 

less ${OUT}/NA12878.highcov.sample.vcf

8. VIEW THE VCF FILES AND COUNT # OF SNPS 

grep -v ^# ${OUT}/NA12878.highcov.sample.vcf | wc -l

9. EVALUATE OVERLAP BETWEEN THE TWO SETS OF VARIANT CALLS 

cat ${OUT}/NA12878.exon.sample.vcf ${OUT}/NA12878.highcov.sample.vcf | grep -v ^# | cut -f 1,2 | sort | uniq -d | wc -l

cat ${OUT}/NA12878.exon.sample.vcf ${OUT}/NA12878.highcov.sample.vcf | grep -v ^# | cut -f 1,2 | sort | uniq -d

10. VIEW ACTUAL ALIGNMENT AT SNP POSITIONS 

${BIN}/samtools-hybrid tview ${OUT}/NA12878.exon.sample.deduped.bam ${REF}/human_g1k_v37_chr20.fa

TYPE g, and 20:19989392 TYPE g, and 20:20032998 TYPE g, and 20:20139952 

${BIN}/samtools-hybrid tview ${IN}/NA12878.highcov.sample.bam ${REF}/human_g1k_v37_chr20.fa

TYPE g, and 20:19989392 TYPE g, and 20:20032998 TYPE g, and 20:20139952

11. SUMMARIZE VCF STATISTICS 

perl ${BIN}/vcfSummary.pl --vcf ${OUT}/NA12878.exon.sample.vcf --dbsnp ${REF}/dbsnp_129_b37.rod.chr20.map --bfile ${REF}/hapmap3_r3_b37_fwd.consensus.qc.poly.chr20

perl ${BIN}/vcfSummary.pl --vcf ${OUT}/NA12878.highcov.sample.vcf --dbsnp ${REF}/dbsnp_129_b37.rod.chr20.map --bfile ${REF}/hapmap3_r3_b37_fwd.consensus.qc.poly.chr20
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