Difference between revisions of "AdvGeneMap2018Commands"
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+ | ===Functional Annotation=== | ||
+ | table_annovar.pl | ||
+ | table_annovar.pl APOC3.vcf humandb/ -buildver hg19 -out APOC3_Gene.vcf -remove -nastring . -protocol refGene -operation g -vcfinput | ||
+ | cat APOC3_Gene.vcf.hg19_multianno.txt | ||
+ | table_annovar.pl APOC3.vcf humandb/ -buildver hg19 -out APOC3_Gene.vcf -remove -nastring . -protocol refGene,knownGene,ensGene -operation g,g,g -arg '-splicing 12 -exonicsplicing','-splicing 12 -exonicsplicing','-splicing 12 -exonicsplicing' -vcfinput | ||
+ | awk -F'\t' '{print $1,$2,$6,$7,$8,$9,$10}' APOC3_Gene.vcf.hg19_multianno.txt | ||
+ | table_annovar.pl APOC3.vcf humandb/ -buildver hg19 -out APOC3_Region.vcf -remove -nastring . -protocol phastConsElements46way -operation r -vcfinput | ||
+ | table_annovar.pl APOC3.vcf humandb/ -buildver hg19 -out APOC3_Region.vcf -remove -nastring . -protocol gwasCatalog -operation r -vcfinput | ||
+ | table_annovar.pl APOC3.vcf humandb/ -buildver hg19 -out APOC3_Filter.vcf -remove -nastring . -protocol gnomad_genome,gnomad_exome,popfreq_max_20150413,gme,avsnp150,dbnsfp33a,dbscsnv11,cadd13gt20,clinvar_20170905,gwava -operation f,f,f,f,f,f,f,f,f,f -vcfinput | ||
+ | awk -F'\t' '{print $1,$2,$103,$104}' APOC3_Filter.vcf.hg19_multianno.txt | ||
+ | awk -F'\t' '{print $1,$2,$6,$14}' APOC3_Filter.vcf.hg19_multianno.txt | ||
+ | awk -F'\t' '{print $1,$2,$15,$16,$17,$18,$19,$20,$21,$22}' APOC3_Filter.vcf.hg19_multianno.txt | ||
+ | awk -F'\t' '{print $1,$2,$36,$86,$70}' APOC3_Filter.vcf.hg19_multianno.txt | ||
+ | awk -F'\t' '{print $1,$2,$99,$100}' APOC3_Filter.vcf.hg19_multianno.txt | ||
+ | table_annovar.pl APOC3.vcf humandb/ -buildver hg19 -out APOC3_ANN.vcf -remove -nastring . -protocol refGene,knownGene,ensGene,wgRna,targetScanS,phastConsElements46way,tfbsConsSites,gwasCatalog,gnomad_genome,gnomad_exome,popfreq_max_20150413,gme,avsnp150,dbnsfp33a,dbscsnv11,cadd13gt20,clinvar_20170905,gwava -operation g,g,g,r,r,r,r,r,f,f,f,f,f,f,f,f,f,f -arg '-splicing 12 -exonicsplicing','-splicing 12 -exonicsplicing','-splicing 12 -exonicsplicing',,,,,,,,,,,,,,, -vcfinput | ||
===GenABEL=== | ===GenABEL=== | ||
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results(testb.qt)$Pc1df[results(testb.qt)$Pc1df < alpha] | results(testb.qt)$Pc1df[results(testb.qt)$Pc1df < alpha] | ||
+ | |||
+ | ===GxG Interaction=== | ||
+ | |||
+ | ./plink --noweb --ped simcasecon.ped --map simcasecon.map --assoc | ||
+ | ./plink --noweb --ped simcasecon.ped --map simcasecon.map --fast-epistasis | ||
+ | ./plink --noweb --ped simcasecon.ped --map simcasecon.map --fast-epistasis --case-only | ||
+ | ./plink --noweb --ped simcasecon.ped --map simcasecon.map --epistasis | ||
+ | ./plink --noweb --ped simcasecon.ped --map simcasecon.map --recodeA --out recoded | ||
+ | ./plink --noweb --ped simcasecon.ped --map simcasecon.map --make-bed --out cassiformat | ||
+ | R | ||
+ | # The following commands are in the R environment | ||
+ | je <-read.table("cassi.out", header=T) | ||
+ | je | ||
+ | library(ORMDR) | ||
+ | recoded<-read.table("recoded.raw", header=T) | ||
+ | head(recoded) | ||
+ | newdata<-recoded[7:106] | ||
+ | ormdrdata<-cbind(newdata,recoded$PHENOTYPE-1) | ||
+ | names(ormdrdata)[101]<-"casestatus" | ||
+ | head(ormdrdata) | ||
+ | mdr1<-mdr.c(ormdrdata, colresp=101, cs=1, combi=1, cv.fold = 10) | ||
+ | mdr1$min.comb | ||
+ | mdr2<-mdr.c(ormdrdata, colresp=101, cs=1, combi=2, cv.fold = 10) | ||
+ | mdr2$min.comb | ||
+ | mdr3<-mdr.c(ormdrdata, colresp=101, cs=1, combi=3, cv.fold = 10) | ||
+ | mdr3$min.comb | ||
+ | mdr1$test.erate | ||
+ | mdr2$test.erate | ||
+ | mdr3$test.erate | ||
+ | mdr1mean<-mean(mdr1$test.erate) | ||
+ | mdr2mean<-mean(mdr2$test.erate) | ||
+ | mdr3mean<-mean(mdr3$test.erate) | ||
+ | mdr1mean | ||
+ | mdr2mean | ||
+ | mdr3mean | ||
+ | mdr2$best.combi | ||
+ | mdr2$min.comb | ||
+ | mdr3$best.combi | ||
+ | mdr3$min.comb | ||
+ | logreg12<-glm(casestatus ~ factor(snp1_2)*factor(snp2_1), family=binomial, | ||
+ | data=ormdrdata) | ||
+ | summary(logreg12) | ||
+ | anova(logreg12) | ||
+ | pchisq(701.68,4,lower.tail=F) | ||
+ | pchisq(703.82,8,lower.tail=F) | ||
+ | logreg345<-glm(casestatus ~ factor(snp3_2)*factor(snp4_2)*factor(snp5_2), | ||
+ | family=binomial, data=ormdrdata) | ||
+ | summary(logreg345) | ||
+ | anova(logreg345) | ||
+ | pchisq(45.6,8,lower.tail=F) | ||
+ | q() | ||
+ | ### The following commands are in the linux shell | ||
+ | ./BEAM3 beam3data.txt -o beam3results | ||
+ | ./BEAM3 beam3data.txt -o beam3results -T 10 | ||
===Plink - Part 1 - Data QC=== | ===Plink - Part 1 - Data QC=== | ||
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#### in R - open R by simply typing R | #### in R - open R by simply typing R | ||
setwd("to_your_working_directory/") | setwd("to_your_working_directory/") | ||
− | sexcheck | + | sexcheck = read.table("GWAS_sex_checking.sexcheck", header=T) |
names(sexcheck) | names(sexcheck) | ||
− | sex_problem | + | sex_problem = sexcheck[which(sexcheck$STATUS=="PROBLEM"),] |
sex_problem | sex_problem | ||
q() | q() | ||
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#### in R | #### in R | ||
setwd("to_your_working_directory/") | setwd("to_your_working_directory/") | ||
− | dups | + | dups = read.table("duplicates.genome", header = T) |
− | problem_pairs | + | problem_pairs = dups[which(dups$PI_HAT > 0.4),] |
problem_pairs | problem_pairs | ||
− | problem_pairs | + | problem_pairs = dups[which(dups$PI_HAT > 0.05),] |
− | myvars | + | myvars = c("FID1", "IID1", "FID2", "IID2", "PI_HAT") |
problem_pairs[myvars] | problem_pairs[myvars] | ||
q() | q() | ||
Line 197: | Line 267: | ||
plink --file GWAS_clean3 --het | plink --file GWAS_clean3 --het | ||
###### in R | ###### in R | ||
− | Dataset <- read.table("plink.het", header | + | Dataset <- read.table("plink.het", header=TRUE, sep="", na.strings="NA", dec=".", strip.white=TRUE) |
mean(Dataset$F) | mean(Dataset$F) | ||
sd(Dataset$F) | sd(Dataset$F) | ||
− | jpeg("hist.jpeg", height | + | jpeg("hist.jpeg", height=1000, width=1000) |
− | hist(scale(Dataset$F), xlim | + | hist(scale(Dataset$F), xlim=c(-4,4)) |
dev.off() | dev.off() | ||
q() | q() | ||
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plink --file GWAS_clean3 --pheno pheno.txt --pheno-name Aff --hardy | plink --file GWAS_clean3 --pheno pheno.txt --pheno-name Aff --hardy | ||
##### in R | ##### in R | ||
− | hardy | + | hardy = read.table("plink.hwe", header = T) |
names(hardy) | names(hardy) | ||
− | hwe_prob | + | hwe_prob = hardy[which(hardy$P < 0.0000009),] |
hwe_prob | hwe_prob | ||
q() | q() | ||
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===Plink - Part 2 - Controlling for Substructure=== | ===Plink - Part 2 - Controlling for Substructure=== | ||
+ | |||
plink --file GWAS_clean4 --genome --cluster --mds-plot 10 | plink --file GWAS_clean4 --genome --cluster --mds-plot 10 | ||
#### in R | #### in R | ||
− | mydata | + | mydata = read.table("mds_components.txt", header=T) |
− | mydata$pch[mydata$Group | + | mydata$pch[mydata$Group==1 ] <-15 |
− | mydata$pch[mydata$Group | + | mydata$pch[mydata$Group==2 ] <-16 |
− | mydata$pch[mydata$Group | + | mydata$pch[mydata$Group==3 ] <-2 |
− | jpeg("mds.jpeg", height | + | jpeg("mds.jpeg", height=500, width=500) |
− | plot(mydata$C1, mydata$C2 ,pch | + | plot(mydata$C1, mydata$C2 ,pch=mydata$pch) |
dev.off() | dev.off() | ||
q() | q() | ||
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broadqq <-function(pvals, title) | broadqq <-function(pvals, title) | ||
{ | { | ||
− | observed <- sort(pvals) | + | observed <- sort(pvals) |
− | lobs <- -(log10(observed)) | + | lobs <- -(log10(observed)) |
− | expected <- c(1:length(observed)) | + | expected <- c(1:length(observed)) |
− | lexp <- -(log10(expected / (length(expected)+1))) | + | lexp <- -(log10(expected / (length(expected)+1))) |
− | plot(c(0,7), c(0,7), col | + | plot(c(0,7), c(0,7), col="red", lwd=3, type="l", xlab="Expected (-logP)", ylab="Observed (-logP)", xlim=c(0,max(lobs)), ylim=c(0,max(lobs)), las=1, xaxs="i", yaxs="i", bty="l", main = title) |
− | points(lexp, lobs, pch | + | points(lexp, lobs, pch=23, cex=.4, bg="black") } |
− | + | jpeg("qqplot_compare.jpeg", height=1000, width=500) | |
− | jpeg("qqplot_compare.jpeg", height | + | par(mfrow=c(2,1)) |
− | par(mfrow | + | aff_unadj<-read.table("unadj.assoc.logistic", header=TRUE) |
− | aff_unadj<-read.table("unadj.assoc.logistic", header | + | aff_unadj.add.p<-aff_unadj[aff_unadj$TEST==c("ADD"),]$P |
− | aff_unadj.add.p<-aff_unadj[aff_unadj$TEST | + | |
broadqq(aff_unadj.add.p,"Some Trait Unadjusted") | broadqq(aff_unadj.add.p,"Some Trait Unadjusted") | ||
− | aff_C1C2<-read.table("PC1-PC2.assoc.logistic", header | + | aff_C1C2<-read.table("PC1-PC2.assoc.logistic", header=TRUE) |
− | aff_C1C2.add.p<-aff_C1C2[aff_C1C2$TEST | + | aff_C1C2.add.p<-aff_C1C2[aff_C1C2$TEST==c("ADD"),]$P |
broadqq(aff_C1C2.add.p, "Some Trait Adjusted for PC1 and PC2") | broadqq(aff_C1C2.add.p, "Some Trait Adjusted for PC1 and PC2") | ||
dev.off() | dev.off() | ||
− | gws_unadj | + | gws_unadj = aff_unadj[which(aff_unadj$P < 0.0000001),] |
gws_unadj | gws_unadj | ||
− | gws_adjusted | + | gws_adjusted = aff_C1C2[which(aff_C1C2$P < 0.0000001),] |
− | gws_adjusted===VAT=== | + | gws_adjusted |
+ | |||
+ | |||
+ | |||
+ | |||
+ | ===RV-TDT=== | ||
+ | ### Variant Annotation | ||
+ | vtools init rvtdt | ||
+ | vtools import --format vcf data/data.vcf --build hg19 | ||
+ | vtools phenotype --from_file data/phen.txt | ||
+ | vtools execute ANNOVAR geneanno | ||
+ | vtools select variant "variant.region_type like '%splicing%'or variant.mut_type like 'nonsynonymous%' or variant.mut_type like 'frameshift%' or variant.mut_type like 'stop%'" -t func_variant | ||
+ | vtools export func_variant --format tped --samples 'phenotype is not null' > vat_raw.tped | ||
+ | # set marker name as chr_pos, needs to avoid duplicate name | ||
+ | sort -k4 -n vat_raw.tped | awk 'BEGIN{OFS="\t";prev="None";copy=1} {$2=$1"_"$4; $3=0; if($2==prev) {$2=$2"_"copy; copy=copy+1} else {prev=$2; copy=1}; print $0}' > vat_export.tped | ||
+ | vtools phenotype --out family sample_name pid mid sex phenotype > vat_export.tfam | ||
+ | vtools use refGene-hg19_20130904 | ||
+ | vtools update func_variant --set 'maf=0.001' # set the maf to be 0.001 | ||
+ | vtools select func_variant -o chr pos refGene.name2 maf --header > vat_export.anno | ||
+ | |||
+ | ### Phasing Trio | ||
+ | plink --noweb --tfile vat_export --recode12 --me 1 1 --set-me-missing --out "recode12_noME" | ||
+ | sort -n -k1 -k6 -k2 recode12_noME.ped | sed 's/ /\t/g' | cut -f1,3,4,5 --complement > linkage.ped cut -f2 recode12_noME.map | awk 'BEGIN{OFS="\t";} {print "M",$0}' | sed '1i\I\tid\nA\tDisease' > linkage.dat | ||
+ | java -Xmx10000m -jar java/linkage2beagle.jar linkage.dat linkage.ped > pre_beagle.bgl | ||
+ | python script/pre_phase.py -i pre_beagle.bgl -a pre_beagle_withMissing.bgl | ||
+ | java -Xmx10000m -jar java/beagle.jar missing=0 trios=pre_beagle.bgl out=bgl_phased verbose=false redundant=true | ||
+ | gunzip bgl_phased.pre_beagle.bgl.phased.gz | ||
+ | |||
+ | ### RV-TDT Analysis | ||
+ | python script/post_phase.py -a vat_export.anno -b bgl_phased.pre_beagle.bgl.phased -o genes/ | ||
+ | for g in `ls genes | grep tped | cut -d"." -f1 | head -20` | ||
+ | do | ||
+ | echo "running rvTDT on gene "${g} | ||
+ | rvTDT exercise_proj -G ./genes/${g}.tped -P ./data/rvtdt.phen -M ./genes/${g}.map --adapt 500 --alpha 0.00001 --permut 2000 --lower_cutoff 0 --upper_cutoff 100 --minVariants 3 --maxMissRatio 1 done | ||
+ | done | ||
+ | |||
+ | |||
+ | |||
+ | ===Seqspark=== | ||
+ | hdfs dfs -put demo.vcf.bz2 | ||
+ | hdfs dfs -put demo.tsv | ||
+ | seqspark annotation.conf | ||
+ | seqspark qc.conf | ||
+ | seqspark demo.conf | ||
+ | |||
+ | ===VAT=== | ||
+ | |||
vtools -h | vtools -h | ||
vtools init VATDemo | vtools init VATDemo |
Latest revision as of 19:46, 23 January 2018
Contents
Functional Annotation
table_annovar.pl table_annovar.pl APOC3.vcf humandb/ -buildver hg19 -out APOC3_Gene.vcf -remove -nastring . -protocol refGene -operation g -vcfinput cat APOC3_Gene.vcf.hg19_multianno.txt table_annovar.pl APOC3.vcf humandb/ -buildver hg19 -out APOC3_Gene.vcf -remove -nastring . -protocol refGene,knownGene,ensGene -operation g,g,g -arg '-splicing 12 -exonicsplicing','-splicing 12 -exonicsplicing','-splicing 12 -exonicsplicing' -vcfinput awk -F'\t' '{print $1,$2,$6,$7,$8,$9,$10}' APOC3_Gene.vcf.hg19_multianno.txt table_annovar.pl APOC3.vcf humandb/ -buildver hg19 -out APOC3_Region.vcf -remove -nastring . -protocol phastConsElements46way -operation r -vcfinput table_annovar.pl APOC3.vcf humandb/ -buildver hg19 -out APOC3_Region.vcf -remove -nastring . -protocol gwasCatalog -operation r -vcfinput table_annovar.pl APOC3.vcf humandb/ -buildver hg19 -out APOC3_Filter.vcf -remove -nastring . -protocol gnomad_genome,gnomad_exome,popfreq_max_20150413,gme,avsnp150,dbnsfp33a,dbscsnv11,cadd13gt20,clinvar_20170905,gwava -operation f,f,f,f,f,f,f,f,f,f -vcfinput awk -F'\t' '{print $1,$2,$103,$104}' APOC3_Filter.vcf.hg19_multianno.txt awk -F'\t' '{print $1,$2,$6,$14}' APOC3_Filter.vcf.hg19_multianno.txt awk -F'\t' '{print $1,$2,$15,$16,$17,$18,$19,$20,$21,$22}' APOC3_Filter.vcf.hg19_multianno.txt awk -F'\t' '{print $1,$2,$36,$86,$70}' APOC3_Filter.vcf.hg19_multianno.txt awk -F'\t' '{print $1,$2,$99,$100}' APOC3_Filter.vcf.hg19_multianno.txt table_annovar.pl APOC3.vcf humandb/ -buildver hg19 -out APOC3_ANN.vcf -remove -nastring . -protocol refGene,knownGene,ensGene,wgRna,targetScanS,phastConsElements46way,tfbsConsSites,gwasCatalog,gnomad_genome,gnomad_exome,popfreq_max_20150413,gme,avsnp150,dbnsfp33a,dbscsnv11,cadd13gt20,clinvar_20170905,gwava -operation g,g,g,r,r,r,r,r,f,f,f,f,f,f,f,f,f,f -arg '-splicing 12 -exonicsplicing','-splicing 12 -exonicsplicing','-splicing 12 -exonicsplicing',,,,,,,,,,,,,,, -vcfinput
GenABEL
# Load files library(GenABEL) convert.snp.tped(tped = "gwa_gabel_qtl.tped", tfam = "gwa_gabel_qtl.tfam", out = "gwa_gabel_qtl.raw", strand = "u") g.dat <- load.gwaa.data(phen = "gwa_gabel_qtl.praw", gen = "gwa_gabel_qtl.raw", force = T) slotNames(g.dat) slotNames(g.dat@gtdata) colnames(g.dat@phdata) # sample size sample.size <- g.dat@gtdata@nids # number of SNPs snps.total <- g.dat@gtdata@nsnps print(c(sample.size, snps.total)) # Trait summary(g.dat@phdata$disease) hist(g.dat@phdata$disease, main="Quantitative Phenotype data summary", xlab = "Systolic pressure measure", freq = F,breaks=20, col="gray") rug(g.dat@phdata$disease) ### # tests for association ### # GLM test test.snp <- scan.glm('disease ~ CRSNP', family = gaussian(), data = g.dat) names(test.snp) alpha <- 5e-8 test.snp$snpnames[test.snp$P1df < alpha] test.snp$P1df[test.snp$P1df < alpha] # Score test test.qt <- qtscore(disease, data = g.dat, trait = "gaussian") slotNames(test.qt) names(test.qt@results) test.qt@lambda descriptives.scan(test.qt) rownames(results(test.qt))[results(test.qt)$P1df < alpha] results(test.qt)$P1df[results(test.qt)$P1df < alpha] results(test.qt)$Pc1df[results(test.qt)$Pc1df < alpha] # QQ plot obs <- sort(results(test.qt)$P1df) ept <- c(1:length(obs)) / (length(obs) + 1) plot(-log10(ept), -log10(obs), main = "GWAS QQ plot, qtl", xlab="Expected -log10(pvalue)", ylab="Observed -log10(pvalue)") abline(0, 1, col = "red") abline(h = 8, lty = 2) # Manhattan plot plot(test.qt, col = "black") # Adding confounders test.qt.sex <- qtscore(disease ~ sex, data = g.dat, trait = "gaussian") rownames(results(test.qt.sex))[results(test.qt)$P1df < alpha] summary(lm(disease ~ sex, data = g.dat)) ### # MDS ### gkin <- ibs(g.dat, weight = "freq") gkin[1:10,1:10] cps.full <- cmdscale(as.dist(.5 - gkin), eig = T, k = 10) names(cps.full) cps <- cps.full$points plot(cps[,1], cps[,2], pch = g.dat@phdata$popn) legend(-0.16, 0.06, c("TSI","MEX", "CEU"), pch = c(1,2,3)) ### # Corrected test ### # Incorporating PCs as predictors colnames(cps)<-c('C1','C2','C3','C4','C5','C6','C7','C8','C9','C10') gpc.dat <- g.dat gpc.dat@phdata<-cbind(g.dat@phdata, cps) test.pc.a <- scan.glm('disease ~ CRSNP + C1 + C2 + C3 + C4 + C5', family=gaussian(), data = gpc.dat) test.pc.a$snpnames[test.pc.a$P1df < alpha] test.pc.a$P1df[test.pc.a$P1df < alpha] test.pc.b <- qtscore(disease ~ C1 + C2 + C3 + C4 + C5, data = gpc.dat, trait = "gaussian") test.pc.b@lambda # scree plot plot(cps.full$eig[1:10]/sum(cps.full$eig), axes = F, type = "b", xlab = "Components", ylim = c(0,0.05), ylab = "Proportion of Variations", main = "MDS analysis scree plot") axis(1, 1:10) axis(2) # cumulative plot plot(cumsum(cps.full$eig[1:10])/sum(cps.full$eig), axes = F, type = "b", ylim = c(0,0.2), xlab = "Components", ylab = "Proportion of Variations", main = "MDS analysis cumulative plot") axis(1, 1:10) axis(2) # Genomic control # Uncorrected GIF test.qt@lambda # Corrected p-value row.names(results(test.qt))[results(test.qt)$Pc1df < alpha] results(test.qt)$Pc1df[results(test.qt)$Pc1df < alpha] # Check for inflation of statistic obs <- sort(results(test.qt)$chi2.1df) ept <- sort(qchisq(1:length(obs) / (length(obs) + 1), df = 1)) plot(ept, obs, main = "Genomic control (slope is the inflation factor)", xlab="Expected chisq, 1df", ylab="Observed chisq, 1df") abline(0, 1, col = "red") abline(0, test.qt@lambda[1], lty = 2) # Definition of GIF # Conventional definition median(results(test.qt)$chi2.1df)/0.456 # GenABEL definition lm(obs~ept)$coef[2] # QQ plot obs <- sort(results(test.qt)$Pc1df) ept <- c(1:length(obs)) / (length(obs) + 1) plot(-log10(ept), -log10(obs), main = "GWAS QQ plot adj. via Genomic Control", xlab="Expected -log10(pvalue)", ylab="Observed -log10(pvalue)") abline(0, 1, col = "red") abline(h = 8, lty = 2) # EIGENSTRAT adj.gkin = gkin diag(adj.gkin) = hom(g.dat)$Var # naxes = 3 is default value test.eg <- egscore(disease, data = g.dat, kin = adj.gkin, naxes = 2) descriptives.scan(test.eg) snp.eg <- row.names(results(test.eg))[results(test.eg)$P1df < alpha] pvalue.eg <- results(test.eg)$P1df[results(test.eg)$P1df < alpha] lambda.eg <- test.eg@lambda snp.eg pvalue.eg lambda.eg # Change #PCs for (k in 1:10){ test.tmp <- egscore(disease, data = g.dat, kin = adj.gkin, naxes = k) print(test.tmp@lambda$estimate) } # QQ plot obs <- sort(results(test.eg)$Pc1df) ept <- c(1:length(obs)) / (length(obs) + 1) qqplot(-log10(ept), -log10(obs), main = "GWAS QQ plot adj. w/ EIGENSTRAT", xlab="Expected -log10(pvalue)", ylab="Observed -log10(pvalue)") abline(0, 1, col = "red") abline(h = 8, lty = 2) # Manhattan plot comparison plot(test.qt, col = "black") add.plot(test.eg, col = "gray", pch = 3) legend("topright", c("Original plot","After correction w/ EIGENSTRAT"), pch = c(1,3)) ### # Basic test, binary trait ### # load files to GenABEL convert.snp.tped(tped = "gwa_gabel.tped", tfam = "gwa_gabel.tfam", out = "gwa_gabel.raw", strand = "u") b.dat <- load.gwaa.data(phen = "gwa_gabel.praw", gen = "gwa_gabel.raw", force = T) slotNames(b.dat) slotNames(b.dat@gtdata) colnames(b.dat@phdata) # sample size b.dat@gtdata@nids # number of cases and controls case.size <- length(which(b.dat@phdata$disease == 1)) control.size <- length(which(b.dat@phdata$disease == 0)) case.size control.size # number of SNPs snpsb.total <- b.dat@gtdata@nsnps # GLM test testb.snp <- scan.glm('disease ~ CRSNP', family = binomial(), data = b.dat) names(testb.snp) alpha <- 5e-8 testb.snp$snpnames[testb.snp$P1df < alpha] testb.snp$P1df[testb.snp$P1df < alpha] # Score test testb.qt <- qtscore(disease, data = b.dat, trait = "binomial") slotNames(testb.qt) descriptives.scan(testb.qt) row.names(results(testb.qt))[results(testb.qt)$P1df < alpha] results(testb.qt)$P1df[results(testb.qt)$P1df < alpha] results(testb.qt)$Pc1df[results(testb.qt)$Pc1df < alpha]
GxG Interaction
./plink --noweb --ped simcasecon.ped --map simcasecon.map --assoc ./plink --noweb --ped simcasecon.ped --map simcasecon.map --fast-epistasis ./plink --noweb --ped simcasecon.ped --map simcasecon.map --fast-epistasis --case-only ./plink --noweb --ped simcasecon.ped --map simcasecon.map --epistasis ./plink --noweb --ped simcasecon.ped --map simcasecon.map --recodeA --out recoded ./plink --noweb --ped simcasecon.ped --map simcasecon.map --make-bed --out cassiformat R # The following commands are in the R environment je <-read.table("cassi.out", header=T) je library(ORMDR) recoded<-read.table("recoded.raw", header=T) head(recoded) newdata<-recoded[7:106] ormdrdata<-cbind(newdata,recoded$PHENOTYPE-1) names(ormdrdata)[101]<-"casestatus" head(ormdrdata) mdr1<-mdr.c(ormdrdata, colresp=101, cs=1, combi=1, cv.fold = 10) mdr1$min.comb mdr2<-mdr.c(ormdrdata, colresp=101, cs=1, combi=2, cv.fold = 10) mdr2$min.comb mdr3<-mdr.c(ormdrdata, colresp=101, cs=1, combi=3, cv.fold = 10) mdr3$min.comb mdr1$test.erate mdr2$test.erate mdr3$test.erate mdr1mean<-mean(mdr1$test.erate) mdr2mean<-mean(mdr2$test.erate) mdr3mean<-mean(mdr3$test.erate) mdr1mean mdr2mean mdr3mean mdr2$best.combi mdr2$min.comb mdr3$best.combi mdr3$min.comb logreg12<-glm(casestatus ~ factor(snp1_2)*factor(snp2_1), family=binomial, data=ormdrdata) summary(logreg12) anova(logreg12) pchisq(701.68,4,lower.tail=F) pchisq(703.82,8,lower.tail=F) logreg345<-glm(casestatus ~ factor(snp3_2)*factor(snp4_2)*factor(snp5_2), family=binomial, data=ormdrdata) summary(logreg345) anova(logreg345) pchisq(45.6,8,lower.tail=F) q() ### The following commands are in the linux shell ./BEAM3 beam3data.txt -o beam3results ./BEAM3 beam3data.txt -o beam3results -T 10
Plink - Part 1 - Data QC
plink --file GWAS plink --file GWAS --mind 0.10 --recode --out GWAS_clean_mind plink --file GWAS_clean_mind --maf 0.05 --recode --out MAF_greater_5 plink --file GWAS_clean_mind --exclude MAF_greater_5.map --recode --out MAF_less_5 plink --file MAF_greater_5 --geno 0.05 --recode --out MAF_greater_5_clean plink --file MAF_less_5 --geno 0.01 --recode --out MAF_less_5_clean plink --file MAF_greater_5_clean --merge MAF_less_5_clean.ped MAF_less_5_clean.map --recode --out GWAS_MAF_clean plink --file GWAS_MAF_clean --mind 0.03 --recode --out GWAS_clean2 plink --file GWAS_clean2 --check-sex --out GWAS_sex_checking #### in R - open R by simply typing R setwd("to_your_working_directory/") sexcheck = read.table("GWAS_sex_checking.sexcheck", header=T) names(sexcheck) sex_problem = sexcheck[which(sexcheck$STATUS=="PROBLEM"),] sex_problem q() ################################## plink --file GWAS_clean2 --genome --out duplicates #### in R setwd("to_your_working_directory/") dups = read.table("duplicates.genome", header = T) problem_pairs = dups[which(dups$PI_HAT > 0.4),] problem_pairs problem_pairs = dups[which(dups$PI_HAT > 0.05),] myvars = c("FID1", "IID1", "FID2", "IID2", "PI_HAT") problem_pairs[myvars] q() ###### plink --file GWAS_clean2 --remove IBS_excluded.txt --recode --out GWAS_clean3 plink --file GWAS_clean3 --het ###### in R Dataset <- read.table("plink.het", header=TRUE, sep="", na.strings="NA", dec=".", strip.white=TRUE) mean(Dataset$F) sd(Dataset$F) jpeg("hist.jpeg", height=1000, width=1000) hist(scale(Dataset$F), xlim=c(-4,4)) dev.off() q() ###### plink --file GWAS_clean3 --pheno pheno.txt --pheno-name Aff --hardy ##### in R hardy = read.table("plink.hwe", header = T) names(hardy) hwe_prob = hardy[which(hardy$P < 0.0000009),] hwe_prob q() ########## plink --file GWAS_clean3 --exclude HWE_out.txt --recode --out GWAS_clean4
Plink - Part 2 - Controlling for Substructure
plink --file GWAS_clean4 --genome --cluster --mds-plot 10 #### in R mydata = read.table("mds_components.txt", header=T) mydata$pch[mydata$Group==1 ] <-15 mydata$pch[mydata$Group==2 ] <-16 mydata$pch[mydata$Group==3 ] <-2 jpeg("mds.jpeg", height=500, width=500) plot(mydata$C1, mydata$C2 ,pch=mydata$pch) dev.off() q() ###### plink --file GWAS_clean4 --pheno pheno.txt --pheno-name Aff --logistic --adjust --out unadj plink --file GWAS_clean4 --genome --cluster --pca 10 header plink --file GWAS_clean4 --pheno pheno.txt --pheno-name Aff --covar plink.eigenvec --covar-name PC1 --logistic --adjust --out PC1 plink --file GWAS_clean4 --pheno pheno.txt --pheno-name Aff --covar plink.eigenvec --covar-name PC1-PC2 --logistic --adjust --out PC1-PC2 #### in R broadqq <-function(pvals, title) { observed <- sort(pvals) lobs <- -(log10(observed)) expected <- c(1:length(observed)) lexp <- -(log10(expected / (length(expected)+1))) plot(c(0,7), c(0,7), col="red", lwd=3, type="l", xlab="Expected (-logP)", ylab="Observed (-logP)", xlim=c(0,max(lobs)), ylim=c(0,max(lobs)), las=1, xaxs="i", yaxs="i", bty="l", main = title) points(lexp, lobs, pch=23, cex=.4, bg="black") } jpeg("qqplot_compare.jpeg", height=1000, width=500) par(mfrow=c(2,1)) aff_unadj<-read.table("unadj.assoc.logistic", header=TRUE) aff_unadj.add.p<-aff_unadj[aff_unadj$TEST==c("ADD"),]$P broadqq(aff_unadj.add.p,"Some Trait Unadjusted") aff_C1C2<-read.table("PC1-PC2.assoc.logistic", header=TRUE) aff_C1C2.add.p<-aff_C1C2[aff_C1C2$TEST==c("ADD"),]$P broadqq(aff_C1C2.add.p, "Some Trait Adjusted for PC1 and PC2") dev.off() gws_unadj = aff_unadj[which(aff_unadj$P < 0.0000001),] gws_unadj gws_adjusted = aff_C1C2[which(aff_C1C2$P < 0.0000001),] gws_adjusted
RV-TDT
### Variant Annotation vtools init rvtdt vtools import --format vcf data/data.vcf --build hg19 vtools phenotype --from_file data/phen.txt vtools execute ANNOVAR geneanno vtools select variant "variant.region_type like '%splicing%'or variant.mut_type like 'nonsynonymous%' or variant.mut_type like 'frameshift%' or variant.mut_type like 'stop%'" -t func_variant vtools export func_variant --format tped --samples 'phenotype is not null' > vat_raw.tped # set marker name as chr_pos, needs to avoid duplicate name sort -k4 -n vat_raw.tped | awk 'BEGIN{OFS="\t";prev="None";copy=1} {$2=$1"_"$4; $3=0; if($2==prev) {$2=$2"_"copy; copy=copy+1} else {prev=$2; copy=1}; print $0}' > vat_export.tped vtools phenotype --out family sample_name pid mid sex phenotype > vat_export.tfam vtools use refGene-hg19_20130904 vtools update func_variant --set 'maf=0.001' # set the maf to be 0.001 vtools select func_variant -o chr pos refGene.name2 maf --header > vat_export.anno ### Phasing Trio plink --noweb --tfile vat_export --recode12 --me 1 1 --set-me-missing --out "recode12_noME" sort -n -k1 -k6 -k2 recode12_noME.ped | sed 's/ /\t/g' | cut -f1,3,4,5 --complement > linkage.ped cut -f2 recode12_noME.map | awk 'BEGIN{OFS="\t";} {print "M",$0}' | sed '1i\I\tid\nA\tDisease' > linkage.dat java -Xmx10000m -jar java/linkage2beagle.jar linkage.dat linkage.ped > pre_beagle.bgl python script/pre_phase.py -i pre_beagle.bgl -a pre_beagle_withMissing.bgl java -Xmx10000m -jar java/beagle.jar missing=0 trios=pre_beagle.bgl out=bgl_phased verbose=false redundant=true gunzip bgl_phased.pre_beagle.bgl.phased.gz ### RV-TDT Analysis python script/post_phase.py -a vat_export.anno -b bgl_phased.pre_beagle.bgl.phased -o genes/ for g in `ls genes | grep tped | cut -d"." -f1 | head -20` do echo "running rvTDT on gene "${g} rvTDT exercise_proj -G ./genes/${g}.tped -P ./data/rvtdt.phen -M ./genes/${g}.map --adapt 500 --alpha 0.00001 --permut 2000 --lower_cutoff 0 --upper_cutoff 100 --minVariants 3 --maxMissRatio 1 done done
Seqspark
hdfs dfs -put demo.vcf.bz2 hdfs dfs -put demo.tsv seqspark annotation.conf seqspark qc.conf seqspark demo.conf
VAT
vtools -h vtools init VATDemo vtools import *.vcf.gz --var_info DP filter --geno_info DP_geno --build hg18 -j1 vtools liftover hg19 head phenotypes.csv vtools phenotype --from_file phenotypes.csv --delimiter "," vtools show project vtools show tables vtools show table variant vtools show samples vtools show genotypes vtools show fields vtools select variant --count vtools show genotypes > GenotypeSummary.txt head GenotypeSummary.txt vtools output variant "max(DP)" "min(DP)" "avg(DP)" "stdev(DP)" "lower_quartile(DP)" "upper_quartile(DP)" --header vtools select variant "filter=’PASS’" --count vtools select variant "filter=’PASS’" -o "max(DP)" "min(DP)" "avg(DP)" "stdev(DP)" "lower_quartile(DP)" "upper_quartile(DP)" --header vtools update variant --from_stat ’total=#(GT)’ ’num=#(alt)’ ’het=#(het)’ ’hom=#(hom)’ ’other=#(other)’ ’minDP=min(DP_geno)’ ’maxDP=max(DP_geno)’ ’meanDP=avg(DP_geno)’ ’maf=maf()’ vtools show fields vtools show table variant vtools update variant --from_stat ’totalGD10=#(GT)’ ’numGD10=#(alt)’ ’hetGD10=#(het)’ ’homGD10=#(hom)’ ’otherGD10=#(other)’ ’mafGD10=maf()’ --genotypes "DP_geno > 10" vtools show fields vtools show table variant vtools output variant chr pos maf mafGD10 --header --limit 20 vtools phenotype --set "RACE=0" --samples "filename like ’YRI%’" vtools phenotype --set "RACE=1" --samples "filename like ’CEU%’" vtools show samples --limit 10 vtools update variant --from_stat ’CEU_mafGD10=maf()’ --genotypes ’DP_geno>10’ --samples "RACE=1" vtools update variant --from_stat ’YRI_mafGD10=maf()’ --genotypes ’DP_geno>10’ --samples "RACE=0" vtools output variant chr pos mafGD10 CEU_mafGD10 YRI_mafGD10 --header --limit 10 vtools phenotype --from_stat ’CEU_totalGD10=#(GT)’ ’CEU_numGD10=#(alt)’ --genotypes ’DP_geno>10’ --samples "RACE=1" vtools phenotype --from_stat ’YRI_totalGD10=#(GT)’ ’YRI_numGD10=#(alt)’ --genotypes ’DP_geno>10’ --samples "RACE=0" vtools phenotype --output sample_nameCEU_totalGD10CEU_numGD10YRI_totalGD10YRI_numGD10 --header vtools execute ANNOVAR geneanno vtools output variant chr pos ref alt mut_type --limit 20 --header vtools_report trans_ratio variant -n num vtools_report trans_ratio variant -n numGD10 vtools select variant "DP<15" -t to_remove vtools show tables vtools remove variants to_remove -v0 vtools show tables vtools remove genotypes "DP_geno<10" -v0 vtools select variant "mut_type like ’non%’ or mut_type like ’stop%’ or region_type=’splicing’" -t v_funct vtools show tables vtools show samples --limit 5 vtools select variant --samples "RACE=1" -t CEU mkdir -p ceu cd ceu vtools init ceu --parent ../ --variants CEU --samples "RACE=1" --build hg19 vtools show project vtools select variant "CEU_mafGD10>=0.05" -t common_ceu vtools select v_funct "CEU_mafGD10<0.01" -t rare_ceu vtools use refGene vtools show annotation refGene vtools associate -h vtools show tests vtools show test LinRegBurden vtools associate common_ceu BMI --covariate SEX -m "LinRegBurden --alternative 2" -j1 --to_db EA_CV > EA_CV.asso.res grep -i error *.log less EA_CV.asso.res sort -g -k7 EA_CV.asso.res | head vtools show fields vtools associate rare_ceu BMI --covariate SEX -m "LinRegBurden --alternative 2" -g refGene.name2 -j1 --to_db EA_RV > EA_RV.asso.res grep -i error *.log | tail -10 less EA_RV.asso.res sort -g -k6 EA_RV.asso.res | head vtools associate rare_ceu BMI --covariate SEX -m "VariableThresholdsQt --alternative 2 -p 100000 --adaptive 0.0005" -g refGene.name2 -j1 --to_db EA_RV > EA_RV_VT.asso.res grep -i error *.log | tail -10 less EA_RV_VT.asso.res sort -g -k6 EA_RV_VT.asso.res | head vtools select rare_ceu "refGene.name2=’ABCC1’" -o chr pos ref alt CEU_mafGD10 numGD10 mut_type --header cd .. vtools select variant --samples "RACE=0" -t YRI mkdir -p yri cd yri vtools init yri --parent ../ --variants YRI --samples "RACE=0" --build hg19 vtools select variant "YRI_mafGD10>=0.05" -t common_yri vtools select v_funct "YRI_mafGD10<0.01" -t rare_yri vtools use refGene vtools associate common_yri BMI --covariate SEX -m "LinRegBurden --alternative 2" -j1 --to_db YA_CV > YA_CV.asso.res vtools associate rare_yri BMI --covariate SEX -m "LinRegBurden --alternative 2" -g refGene.name2 -j1 --to_db YA_RV > YA_RV.asso.res vtools associate rare_yri BMI --covariate SEX -m "VariableThresholdsQt --alternative 2 -p 100000 --adaptive 0.0005" -g refGene.name2 -j1 --to_db YA_RV > YA_RV_VT.asso.res cd .. vtools_report meta_analysis ceu/EA_RV_VT.asso.res yri/YA_RV_VT.asso.res --beta 5 --pval 6 --se 7 -n 2 --link 1 > ME\ TA_RV_VT.asso.res cut -f1,3 META_RV_VT.asso.res | head