Last updated: 2020-06-17

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Knit directory: apaQTL/analysis/

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Unstaged changes:
    Modified:   analysis/NuclearSpecIncludeNotTested.Rmd
    Modified:   analysis/PASdescriptiveplots.Rmd
    Modified:   analysis/Readdistagainstfeatures.Rmd
    Modified:   analysis/TSS.Rmd
    Modified:   analysis/apabyeQTLstatus.Rmd
    Modified:   analysis/decayAndStability.Rmd
    Modified:   analysis/miRNAdisrupt.Rmd
    Modified:   analysis/nascenttranscription.Rmd
    Modified:   analysis/nucSpecinEQTLs.Rmd
    Modified:   analysis/overlapapaqtlsandeqtls.Rmd
    Modified:   analysis/pQTLexampleplot.Rmd
    Modified:   analysis/version15bpfilter.Rmd
    Modified:   code/DistPAS2Sig.py
    Modified:   code/Script4NuclearQTLexamples.sh
    Modified:   code/Script4TotalQTLexamples.sh
    Modified:   code/apaQTLsnake.err
    Modified:   code/apaqtlfacetboxplots.R
    Modified:   code/environment.yaml
    Modified:   code/run_qtlFacetBoxplots.sh
    Deleted:    code/test.txt
    Deleted:    reads_graphs.Rmd

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These are the previous versions of the R Markdown and HTML files. If you’ve configured a remote Git repository (see ?wflow_git_remote), click on the hyperlinks in the table below to view them.

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Rmd a69cf8d brimittleman 2020-06-17 add 1-1 line
html 85007a7 brimittleman 2019-09-07 Build site.
Rmd 3088c20 brimittleman 2019-09-07 add tot spec PAS
html 7711ef9 brimittleman 2019-09-04 Build site.
Rmd 73f5833 brimittleman 2019-09-04 wflow_publish(c(“analysis/QTLlocation.Rmd”, “analysis/apaQTLoverlap.Rmd”, “analysis/nonNormQTL.Rmd”,
html 3875f36 brimittleman 2019-08-01 Build site.
Rmd 3d49e9f brimittleman 2019-08-01 figure2D
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Rmd 4fd345d brimittleman 2019-08-01 figure2D
html 6757ce6 brimittleman 2019-07-17 Build site.
Rmd bf42ee6 brimittleman 2019-07-17 remove density
html 4297ae4 brimittleman 2019-07-03 Build site.
Rmd 2cafaf1 brimittleman 2019-07-03 include spec qtl
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Rmd 968cb4b brimittleman 2019-06-13 wflow_publish(“analysis/nonNormQTL.Rmd”)
html 230bc6a brimittleman 2019-06-11 Build site.
Rmd cd24c16 brimittleman 2019-06-11 new genos
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Rmd 6ece235 brimittleman 2019-05-28 add example code
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Rmd c04929f brimittleman 2019-05-28 add example code
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Rmd f10e64d brimittleman 2019-05-28 add plot grid
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Rmd f5260bd brimittleman 2019-05-28 add results
html 28c8ca3 brimittleman 2019-05-24 Build site.
Rmd 3f63045 brimittleman 2019-05-24 add code to prepare non norm qtl

In order to compare effect sizes for the QTLs I have previously identified in an interpretable manner, I need to run the linear model with the non normalized usage. To do this I will separate the the usage (with annotation) files by chromosome and run fastqtl on these files.

library(workflowr)
This is workflowr version 1.6.0
Run ?workflowr for help getting started
library(tidyverse)
── Attaching packages ───────────────────────────────────── tidyverse 1.2.1 ──
✔ ggplot2 3.1.1       ✔ purrr   0.3.2  
✔ tibble  2.1.1       ✔ dplyr   0.8.0.1
✔ tidyr   0.8.3       ✔ stringr 1.3.1  
✔ readr   1.3.1       ✔ forcats 0.3.0  
── Conflicts ──────────────────────────────────────── tidyverse_conflicts() ──
✖ dplyr::filter() masks stats::filter()
✖ dplyr::lag()    masks stats::lag()
library(cowplot)

Attaching package: 'cowplot'
The following object is masked from 'package:ggplot2':

    ggsave

Prepare files

countsnum= APApeak_Phenotype_GeneLocAnno.Total.5perc.CountsNumeric, APApeak_Phenotype_GeneLocAnno.Nuclear.5perc.CountsNumeric

id file= APApeak_Phenotype_GeneLocAnno.Nuclear.5perc.fc.gz, APApeak_Phenotype_GeneLocAnno.Total.5perc.fc.gz

totAnno= read.table("../data/phenotype_5perc/APApeak_Phenotype_GeneLocAnno.Total.5perc.fc.gz", stringsAsFactors = F, header = T) %>% separate(chrom, into=c("Chrchrom", "Start", "End", "ID"),sep=":") %>% mutate(Chr=str_sub(Chrchrom, 4, str_length(Chrchrom)))
                                                                                                                                                                                                                
colnamesTot= colnames(totAnno)[5:56]
totUsage=read.table("../data/phenotype_5perc/APApeak_Phenotype_GeneLocAnno.Total.5perc.CountsNumeric", stringsAsFactors = F, header = F, col.names = colnamesTot) 

totUsageAnno=as.data.frame(cbind(Chr=totAnno$Chr, start=totAnno$Start, end=totAnno$End, ID=totAnno$ID, totUsage ))

write.table(totUsageAnno,file="../data/nonNorm_pheno/TotalUsageAllChrom.txt", col.names = T, row.names = F, quote = F, sep="\t" )
nucAnno= read.table("../data/phenotype_5perc/APApeak_Phenotype_GeneLocAnno.Nuclear.5perc.fc.gz", stringsAsFactors = F, header = T)%>% separate(chrom, into=c("Chrchrom", "Start", "End", "ID"),sep=":") %>% mutate(Chr=str_sub(Chrchrom, 4, str_length(Chrchrom)))
colnamesNuc= colnames(nucAnno)[5:56]
nucUsage=read.table("../data/phenotype_5perc/APApeak_Phenotype_GeneLocAnno.Nuclear.5perc.CountsNumeric", stringsAsFactors = F, header = F, col.names = colnamesNuc) 


nucUsageAnno=as.data.frame(cbind(Chr=nucAnno$Chr, start=nucAnno$Start, end=nucAnno$End, ID=nucAnno$ID, nucUsage ))

write.table(nucUsageAnno,file="../data/nonNorm_pheno/NuclearUsageAllChrom.txt", col.names = T, row.names = F, quote = F, sep="\t" )

Run QTL scripts

I will create a python script to seperate the file into each chromosome for running fastQTL.

sbatch run_sepUsagephen.sh
sbatch ZipandTabPheno.sh
sbatch ApaQTL_nominalNonnorm.sh

Concatinate files:

cat  ../data/nonNorm_pheno/TotalUsageChrom*.nominal.out > ../data/nonNorm_pheno/TotalUsageChrom_Nominal_AllChrom.txt
cat ../data/nonNorm_pheno/NuclearUsageChrom*.nominal.out > ../data/nonNorm_pheno/NuclearUsageChrom_Nominal_AllChrom.txt

Pull out real total and nuc QLTs

python qtlsPvalOppFrac.py   ../data/apaQTLs/Nuclear_apaQTLs4pc_5fdr.txt ../data/nonNorm_pheno/TotalUsageChrom_Nominal_AllChrom.txt ../data/QTLoverlap_nonNorm/NuclearQTLinTotalNominal_nonNorm.txt  

python qtlsPvalOppFrac.py ../data/apaQTLs/Nuclear_apaQTLs4pc_5fdr.txt ../data/nonNorm_pheno/NuclearUsageChrom_Nominal_AllChrom.txt ../data/QTLoverlap_nonNorm/NuclearQTLinNuclearNominal_nonNorm.txt  


python qtlsPvalOppFrac.py ../data/apaQTLs/Total_apaQTLs4pc_5fdr.txt ../data/nonNorm_pheno/TotalUsageChrom_Nominal_AllChrom.txt ../data/QTLoverlap_nonNorm/TotalQTLinTotalNominal_nonNorm.txt  

python qtlsPvalOppFrac.py ../data/apaQTLs/Total_apaQTLs4pc_5fdr.txt ../data/nonNorm_pheno/NuclearUsageChrom_Nominal_AllChrom.txt ../data/QTLoverlap_nonNorm/TotalQTLinNuclearNominal_nonNorm.txt  
totAPAinNuc=read.table("../data/QTLoverlap_nonNorm/TotalQTLinNuclearNominal_nonNorm.txt", header = F, stringsAsFactors = F, col.names=c("peakID", "snp", "dist", "pval", "slope"))


nucAPAinTot=read.table("../data/QTLoverlap_nonNorm/NuclearQTLinTotalNominal_nonNorm.txt", header = F, stringsAsFactors = F, col.names=c("peakID", "snp", "dist", "pval", "slope"))

totAPAinTot=read.table("../data/QTLoverlap_nonNorm/TotalQTLinTotalNominal_nonNorm.txt", header = F, stringsAsFactors = F, col.names=c("peakID", "snp", "dist", "pval", "slope")) %>% dplyr::select(peakID, snp, slope) %>% dplyr::rename("Originalslope"=slope)

nucAPAinNuc=read.table("../data/QTLoverlap_nonNorm/NuclearQTLinNuclearNominal_nonNorm.txt", header = F, stringsAsFactors = F, col.names=c("peakID", "snp", "dist", "pval", "slope")) %>% dplyr::select(peakID, snp, slope)%>% dplyr::rename("Originalslope"=slope)

Total

TotBoth= totAPAinNuc %>% inner_join(totAPAinTot,by=c("peakID", "snp"))

summary(lm(TotBoth$slope ~ TotBoth$Originalslope))

Call:
lm(formula = TotBoth$slope ~ TotBoth$Originalslope)

Residuals:
     Min       1Q   Median       3Q      Max 
-0.66101 -0.03117  0.00361  0.03873  0.37205 

Coefficients:
                       Estimate Std. Error t value Pr(>|t|)    
(Intercept)           -0.004458   0.004247   -1.05    0.295    
TotBoth$Originalslope  0.638860   0.022554   28.33   <2e-16 ***
---
Signif. codes:  0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1

Residual standard error: 0.0852 on 401 degrees of freedom
Multiple R-squared:  0.6668,    Adjusted R-squared:  0.6659 
F-statistic: 802.4 on 1 and 401 DF,  p-value: < 2.2e-16
totbothplot=ggplot(TotBoth, aes(x=Originalslope, y=slope))+geom_point() + geom_smooth(method="lm") + labs(title="Total apaQTL effect sizes", x="Effect size in Total",y="Effect size in Nucler") + geom_density_2d(col="red") + annotate("text", y=1, x=0, label="R2=.67, slope=0.64")

Nuclear

NucBoth= nucAPAinTot %>% inner_join(nucAPAinNuc,by=c("peakID", "snp"))
summary(lm(NucBoth$slope ~ NucBoth$Originalslope))

Call:
lm(formula = NucBoth$slope ~ NucBoth$Originalslope)

Residuals:
     Min       1Q   Median       3Q      Max 
-0.23354 -0.03332  0.00028  0.03182  0.40042 

Coefficients:
                      Estimate Std. Error t value Pr(>|t|)    
(Intercept)           0.001628   0.002910   0.559    0.576    
NucBoth$Originalslope 0.775971   0.017539  44.243   <2e-16 ***
---
Signif. codes:  0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1

Residual standard error: 0.06408 on 483 degrees of freedom
Multiple R-squared:  0.8021,    Adjusted R-squared:  0.8017 
F-statistic:  1957 on 1 and 483 DF,  p-value: < 2.2e-16
Nucbothplot=ggplot(NucBoth, aes(x=Originalslope, y=slope))+geom_point() + geom_smooth(method="lm") + labs(title="Nuclear apaQTL effect sizes", x="Effect size in Nuclear",y="Effect size in Total") + geom_density_2d(col="red") +  annotate("text", y=1, x=0, label="R2=.8, slope=0.78")
plot_grid(totbothplot,Nucbothplot)

Version Author Date
7711ef9 brimittleman 2019-09-04
0c7c682 brimittleman 2019-06-13
230bc6a brimittleman 2019-06-11
de2aa7e brimittleman 2019-05-28
f4a2106 brimittleman 2019-05-28

For the nuclear plot I want to include the PAS we cannot test in the total fraction. To do this I will write code that only writes out the lines for the PAS not in the total fraction. I need the nominal effect sizes for the nuclear qtls in PAS we could not test in total. First I will get all of the PAS tested in total and figure out the nuclear QTLs not in this set. I will then pull those associations out of the nominal nuclear set.

python nucSpeceffectsize.py
nucSpec=read.table("../data/QTLoverlap_nonNorm/NuclearSpecQTLinNuclearNominal_nonNorm.txt",header = F, stringsAsFactors = F, col.names=c("peakID", "snp", "dist", "pval", "Originalslope")) %>% mutate(slope=0,set="Specific") %>% select(peakID, snp, dist, pval, Originalslope, slope,set)
NucBoth_set= NucBoth %>% mutate(set="Both")
NucBothwSpec=bind_rows(NucBoth_set, nucSpec)
summary(lm(NucBothwSpec$slope ~ NucBothwSpec$Originalslope))

Call:
lm(formula = NucBothwSpec$slope ~ NucBothwSpec$Originalslope)

Residuals:
     Min       1Q   Median       3Q      Max 
-0.29479 -0.03377  0.00364  0.03780  0.34739 

Coefficients:
                            Estimate Std. Error t value Pr(>|t|)    
(Intercept)                -0.009236   0.002974  -3.106  0.00199 ** 
NucBothwSpec$Originalslope  0.664058   0.018388  36.114  < 2e-16 ***
---
Signif. codes:  0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1

Residual standard error: 0.07255 on 600 degrees of freedom
Multiple R-squared:  0.6849,    Adjusted R-squared:  0.6844 
F-statistic:  1304 on 1 and 600 DF,  p-value: < 2.2e-16
cor.test(y=NucBothwSpec$slope, x=NucBothwSpec$Originalslope)$p.value
[1] 1.336923e-152
nucspecplot=ggplot(NucBothwSpec, aes(x=Originalslope, y=slope))+geom_point(aes( col=set)) + geom_smooth(method="lm") + labs(title="Nuclear apaQTL effect sizes", x="Effect size in Nuclear",y="Effect size in Total") +  annotate("text", y=1, x=0, label="R2=.68, slope=0.66",size=6) + scale_color_brewer(palette = "Dark2")+ theme(text = element_text(size=16), legend.position = "bottom")

Do this in the other fraction to see what is looks like.

python totSeceffectsize.py
totSpec=read.table("../data/QTLoverlap_nonNorm/TotalSpecQTLinTotalNominal_nonNorm.txt",header = F, stringsAsFactors = F, col.names=c("peakID", "snp", "dist", "pval", "Originalslope")) %>% mutate(slope=0,set="Specific") %>% select(peakID, snp, dist, pval, Originalslope, slope,set)
TotBoth_set= TotBoth %>% mutate(set="Both")
TotBothwSpec=bind_rows(TotBoth_set, totSpec)
summary(lm(TotBothwSpec$slope ~ TotBothwSpec$Originalslope))

Call:
lm(formula = TotBothwSpec$slope ~ TotBothwSpec$Originalslope)

Residuals:
     Min       1Q   Median       3Q      Max 
-0.64645 -0.03081  0.00337  0.03800  0.37917 

Coefficients:
                            Estimate Std. Error t value Pr(>|t|)    
(Intercept)                -0.006902   0.003969  -1.739   0.0827 .  
TotBothwSpec$Originalslope  0.621773   0.021776  28.553   <2e-16 ***
---
Signif. codes:  0 '***' 0.001 '**' 0.01 '*' 0.05 '.' 0.1 ' ' 1

Residual standard error: 0.08339 on 441 degrees of freedom
Multiple R-squared:  0.649, Adjusted R-squared:  0.6482 
F-statistic: 815.2 on 1 and 441 DF,  p-value: < 2.2e-16
totspecplot=ggplot(TotBothwSpec, aes(x=Originalslope, y=slope))+geom_point(aes( col=set)) + geom_smooth(method="lm") + labs(title="Total apaQTL effect sizes", x="Effect size in Total",y="Effect size in Nuclear") +  annotate("text", y=1, x=0, label="R2=.65, slope=0.62", size=6) + scale_color_brewer(palette = "Dark2") + theme(text = element_text(size=16), legend.position = "bottom")

totspecplot

Version Author Date
7711ef9 brimittleman 2019-09-04
6757ce6 brimittleman 2019-07-17
grideffectplot=plot_grid(nucspecplot,totspecplot)
grideffectplot

Version Author Date
7711ef9 brimittleman 2019-09-04
6757ce6 brimittleman 2019-07-17
4297ae4 brimittleman 2019-07-03
NucBothwSpec_onlyboth=NucBothwSpec %>% filter(set=="Both")
nucspecplot2=ggplot(NucBothwSpec_onlyboth, aes(x=Originalslope, y=slope))+geom_point(col="#1B9E77") + geom_smooth(method="lm") + labs( x="Effect size in Nuclear",y="Effect size in Total") + theme(text = element_text(size=16), legend.position = "bottom") + geom_abline(slope = 1)

nucspecplot2

TotBothwSpec %>% filter(set=="Specific") %>% nrow()
[1] 40
NucBothwSpec %>% filter(set=="Specific") %>% nrow()
[1] 117
prop.test(x=c(117,40), n=c(565,378), alternative ="greater")

    2-sample test for equality of proportions with continuity
    correction

data:  c(117, 40) out of c(565, 378)
X-squared = 16.012, df = 1, p-value = 3.147e-05
alternative hypothesis: greater
95 percent confidence interval:
 0.06079568 1.00000000
sample estimates:
   prop 1    prop 2 
0.2070796 0.1058201 

specific PAS:

totalPAS=read.table("../data/PAS/APApeak_Peaks_GeneLocAnno.Total.5perc.sort.bed",stringsAsFactors = F,col.names = c("chr", "start","end","name", "score", "strand"))
nuclearPAS=read.table("../data/PAS/APApeak_Peaks_GeneLocAnno.Nuclear.5perc.sort.bed",stringsAsFactors = F,col.names = c("chr", "start","end","name", "score", "strand"))

totalSpec=totalPAS %>% anti_join(nuclearPAS,by="name")
40/nrow(totalSpec)
[1] 0.01509434

sessionInfo()
R version 3.5.1 (2018-07-02)
Platform: x86_64-pc-linux-gnu (64-bit)
Running under: Scientific Linux 7.4 (Nitrogen)

Matrix products: default
BLAS/LAPACK: /software/openblas-0.2.19-el7-x86_64/lib/libopenblas_haswellp-r0.2.19.so

locale:
 [1] LC_CTYPE=en_US.UTF-8       LC_NUMERIC=C              
 [3] LC_TIME=en_US.UTF-8        LC_COLLATE=en_US.UTF-8    
 [5] LC_MONETARY=en_US.UTF-8    LC_MESSAGES=en_US.UTF-8   
 [7] LC_PAPER=en_US.UTF-8       LC_NAME=C                 
 [9] LC_ADDRESS=C               LC_TELEPHONE=C            
[11] LC_MEASUREMENT=en_US.UTF-8 LC_IDENTIFICATION=C       

attached base packages:
[1] stats     graphics  grDevices utils     datasets  methods   base     

other attached packages:
 [1] cowplot_0.9.4   forcats_0.3.0   stringr_1.3.1   dplyr_0.8.0.1  
 [5] purrr_0.3.2     readr_1.3.1     tidyr_0.8.3     tibble_2.1.1   
 [9] ggplot2_3.1.1   tidyverse_1.2.1 workflowr_1.6.0

loaded via a namespace (and not attached):
 [1] tidyselect_0.2.5   haven_1.1.2        lattice_0.20-38   
 [4] colorspace_1.3-2   generics_0.0.2     htmltools_0.3.6   
 [7] yaml_2.2.0         rlang_0.4.0        later_0.7.5       
[10] pillar_1.3.1       glue_1.3.0         withr_2.1.2       
[13] RColorBrewer_1.1-2 modelr_0.1.2       readxl_1.1.0      
[16] plyr_1.8.4         munsell_0.5.0      gtable_0.2.0      
[19] cellranger_1.1.0   rvest_0.3.2        evaluate_0.12     
[22] labeling_0.3       knitr_1.20         httpuv_1.4.5      
[25] broom_0.5.1        Rcpp_1.0.4.6       promises_1.0.1    
[28] scales_1.0.0       backports_1.1.2    jsonlite_1.6      
[31] fs_1.3.1           hms_0.4.2          digest_0.6.18     
[34] stringi_1.2.4      grid_3.5.1         rprojroot_1.3-2   
[37] cli_1.1.0          tools_3.5.1        magrittr_1.5      
[40] lazyeval_0.2.1     crayon_1.3.4       whisker_0.3-2     
[43] pkgconfig_2.0.2    MASS_7.3-51.1      xml2_1.2.0        
[46] lubridate_1.7.4    assertthat_0.2.0   rmarkdown_1.10    
[49] httr_1.3.1         rstudioapi_0.10    R6_2.3.0          
[52] nlme_3.1-137       git2r_0.26.1       compiler_3.5.1