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| exit PuTTY | | exit PuTTY |
| | | |
− | === Tuesday FEEDBACK! ===
| |
− | Please provide feedback on the lectures/tutorials from today:
| |
− |
| |
− | https://docs.google.com/forms/d/1n8xYxvsOq-HsabpDfGcHvwD84BYIRDx8_b-H5N3d-D8/viewform
| |
| </div> | | </div> |
| </div> | | </div> |
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| <div class="mw-collapsible mw-collapsed" style="width:500px"> | | <div class="mw-collapsible mw-collapsed" style="width:500px"> |
| + | |
| == Wednesday == | | == Wednesday == |
| <div class="mw-collapsible-content"> | | <div class="mw-collapsible-content"> |
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| exit PuTTY | | exit PuTTY |
| | | |
− | ==== Wednesday FEEDBACK! ====
| |
− | Please provide feedback on the lectures/tutorials from today:
| |
− |
| |
− | https://docs.google.com/a/umich.edu/forms/d/1CCHL9ODPsw4jX4hj0kGo6AMHwT4Gam0IpKNRnIR9yMk/viewform
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| | | |
| </div> | | </div> |
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| <div class="mw-collapsible mw-collapsed" style="width:500px"> | | <div class="mw-collapsible mw-collapsed" style="width:500px"> |
| + | |
| == Thursday == | | == Thursday == |
| <div class="mw-collapsible-content"> | | <div class="mw-collapsible-content"> |
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| </div> | | </div> |
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− | <div class="mw-collapsible" style="width:500px"> | + | <div class="mw-collapsible" style="width:1000px"> |
| | | |
| == Friday == | | == Friday == |
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| Look at the vcfs: | | Look at the vcfs: |
| ls ~/$SAMPLE/output/vcfs | | ls ~/$SAMPLE/output/vcfs |
| + | |
| + | === Friday : More SNP Analysis === |
| + | |
| + | ==== Environmental Variables ==== |
| + | |
| + | If you didn't set the environmental variable, you can set it again |
| + | |
| + | source /net/seqshop-server/home/mktrost/seqshop/setup.txt |
| + | export SAMPLE=SampleXX (MAKE SURE TO CHANGE XX to your number or use NA12878 instead) |
| + | source /net/seqshop-server/home/mktrost/seqshop/setupSS.txt |
| + | |
| + | In addition, set another environmental variable for locating the binaries for custom analysis |
| + | |
| + | export HK=/net/seqshop-server/home/hmkang/seqshop/bin |
| + | |
| + | ==== Annotation / Lookup against dbSNP ==== |
| + | |
| + | If you want to add rsIDs to your variant files, you can do this by running the following command |
| + | |
| + | $HK/vcf-add-rsid -vcf $OUT/vcfs/chr1/chr1.filtered.vcf.gz --db $HK/../data/dbSNP.b138/dbsnp_138.b37.vcf.gz --out $OUT/vcfs/chr1/chr1.filtered.rsid.vcf.gz |
| + | |
| + | If you want to run this command across all chromosomes in parallel, you can use the special script run-command-wgs |
| + | |
| + | $HK/run-command-wgs --cmd "$HK/vcf-add-rsid -vcf $OUT/vcfs/chr1/chr1.filtered.vcf.gz --db $HK/../data/dbSNP.b138/dbsnp_138.b37.vcf.gz --out $OUT/vcfs/chr1/chr1.filtered.rsid.vcf.gz" --numjobs 6 |
| + | |
| + | Looking up SNPs by rsID is possible by (for example) |
| + | $HK/vcf-lookup-rsid --vcf $OUT/vcfs/chr1/chr1.filtered.vcf.gz --sepchr --rs rs17766217 |
| + | * Be sure to look at the QUAL & your sample's PL, and not just the GL field. Check if QUAL is 0 or PL is 0,0,0 - NS is also probably 0; DP is probably 0. That means you probably didn't have any copies, so your GT may not be correct/is unknown. |
| + | |
| + | If you want to browse the rsIDs of known GWAS SNPs, you can do this by |
| + | cut -f 1,8,22 $HK/../data/gwascatalog/gwascatalog.txt | less |
| + | |
| + | ==== Annotating your genome ==== |
| + | |
| + | You can annotate your genome using EPACTS software packages. Individual chromosome can be annotated by running. |
| + | $EPACTS/bin/epacts anno --in $OUT/vcfs/chr1/chr1.filtered.rsid.vcf.gz --out $OUT/vcfs/chr1/chr1.filtered.rsid.anno.vcf.gz |
| + | |
| + | Or you can run multiple chromosomes in parallel in one command |
| + | $HK/run-command-wgs --cmd "$EPACTS/bin/epacts anno --in $OUT/vcfs/chr1/chr1.filtered.rsid.vcf.gz --out $OUT/vcfs/chr1/chr1.filtered.rsid.anno.vcf.gz" --numjobs 6 |
| + | |
| + | ==== Extracting only exonic SNPs ==== |
| + | |
| + | If you want to look at the exonic SNPs, you can extract using the following command |
| + | $HK/run-command-wgs --cmd "($HK/tabix -H $OUT/vcfs/chr1/chr1.filtered.rsid.anno.vcf.gz; zcat $OUT/vcfs/chr1/chr1.filtered.rsid.anno.vcf.gz | grep Exon;)| $HK/bgzip -c > $OUT/vcfs/chr1/chr1.filtered.rsid.anno.exon.vcf.gz" --numjobs 6 |
| + | |
| + | And they can be combined as follows |
| + | (zcat $OUT/vcfs/chr1/chr1.filtered.rsid.anno.exon.vcf.gz; zcat $OUT/vcfs/chr[2-9]/chr*.filtered.rsid.anno.exon.vcf.gz $OUT/vcfs/chr??/chr*.filtered.rsid.anno.exon.vcf.gz $OUT/vcfs/chrX/chrX.filtered.rsid.anno.exon.vcf.gz | grep -v ^#) | $HK/bgzip -c > $OUT/wgs.filtered.rsid.anno.exon.vcf.gz |
| + | |
| + | ==== Exonic Variants NOT found by 1000G ==== |
| + | |
| + | If you are interested in rare variants that are not identified by 1000G, you can extract them by running |
| + | zcat $OUT/wgs.filtered.rsid.anno.exon.vcf.gz | grep "EXTFILTER=NA,NA" | grep -v -w "0/0" | less |
| + | |
| + | For example, |
| + | zcat $OUT/wgs.filtered.rsid.anno.exon.vcf.gz | grep "EXTFILTER=NA,NA" | grep -v -w "0/0" | perl -lane 'print "$1\t$F[6]" if ( /ANNO=([^;:]+)/)' | sort | uniq -c |
| + | |
| + | will give you the counts of variants, separate by the filtering results |
| + | |
| + | * Q1. How manny novel silent, missense, and nonsense SNPs are found? Is that too few, too small, or just about right? |
| + | * Q2. Looking at each functional category, which functional categories has largest fraction of SNPs failed filter? Why do you think it is? |
| + | * Q3. Can you exclude the sites that are also in dbSNP, and count how many nonsense variants are left? |
| + | |
| + | |
| + | To also exclude those in dbsnp: |
| + | zcat $OUT/wgs.filtered.rsid.anno.exon.vcf.gz | grep "EXTFILTER=NA,NA" | grep -v -w "0/0" | grep -v rs| perl -lane 'print "$1\t$F[6]" if ( /ANNO=([^;:]+)/)' | sort | uniq -c |
| + | |
| + | Exclude dbsnp and look at Stop_Gain variants |
| + | zcat $OUT/wgs.filtered.rsid.anno.exon.vcf.gz | grep "EXTFILTER=NA,NA" | grep -v -w "0/0" |grep -v rs | perl -lane 'print "$_" if ( /ANNO=Stop_Gain/)' |grep -w PASS |
| + | |
| + | Want to see this from the BAM file? Use samtools tview: |
| + | $GC/bin/samtools tview $SAMPLE/output/bams/$SAMPLE.recal.bam $GC/gotcloud.ref/human.g1k.v37.fa |
| + | Use 'g' & enter the Chr:Pos |
| + | * Some patterns may indicate not real variants. |
| + | |
| + | If you want to know predicted functional significance of a particular variant, you can search by |
| + | |
| + | $HK/tabix $HK/../data/CADD/whole_genome_SNVs.tsv.gz [chr]:[pos] | head -3 |
| + | |
| + | The phred score at the last column quantifies the degree of functional significance |
| | | |
| | | |
− | ==== WORKSHOP FEEDBACK! ====
| |
− | Please provide feedback on the workshop in general:
| |
| | | |
− | https://docs.google.com/forms/d/1f8HjTKvxgYuApl9dLNqbW9Su3N3v9jtMpEDcbPJ1PYk/viewform
| |
| </div> | | </div> |
| </div> | | </div> |