# HG changeset patch # User joachim-jacob # Date 1374048372 14400 # Node ID a48dde9abecf0d36667343764b5e8210da19afa1 Uploaded initial version diff -r 000000000000 -r a48dde9abecf README --- /dev/null Thu Jan 01 00:00:00 1970 +0000 +++ b/README Wed Jul 17 04:06:12 2013 -0400 @@ -0,0 +1,28 @@ +VSTransformation - 1.0.0 +============================= + + +SUMMARY +------- +This tool takes in a count table generated from a mapping of RNA-seq data, +and performs a variance stabilizing transformation on these counts. + +REQUIREMENTS AND INSTALLATION +----------------------------- +Install this tool in your Galaxy via the Toolshed + + ) Third party code installation + You need to have a working R environment available in your path, + with following dependencies installed: + - reshape + - ggplot2 + - deseq + - knitr + - markdown + + ) Integration into Galaxy + Just use the Management interface to integrate this tool. + +DETAILS AND ISSUES +------------------ + ) Soon dependencies will be resolved. diff -r 000000000000 -r a48dde9abecf test-data/counttable1.tsv --- /dev/null Thu Jan 01 00:00:00 1970 +0000 +++ b/test-data/counttable1.tsv Wed Jul 17 04:06:12 2013 -0400 @@ -0,0 +1,15683 @@ +GeneID can_1 can_2 nbg_1 nbg_2 n18_1 n18_2 n25_1 n25_2 +FBgn0000003 33 41 150 1817 35 58 168 68 +FBgn0000008 798 711 630 777 679 721 626 792 +FBgn0000014 0 3 1 1 3 3 3 5 +FBgn0000015 0 0 1 0 2 0 1 0 +FBgn0000017 4335 3393 3201 4123 3603 3891 3254 4085 +FBgn0000018 165 142 138 135 116 147 112 174 +FBgn0000022 0 0 0 0 0 0 0 0 +FBgn0000024 7241 5601 4691 6767 5270 6021 5116 6197 +FBgn0000028 356 307 293 378 381 442 336 413 +FBgn0000032 318 295 267 304 320 342 277 331 +FBgn0000036 845 588 421 761 708 760 597 766 +FBgn0000037 3167 2600 2629 3092 2138 2397 2126 2751 +FBgn0000038 16133 12965 13243 14216 11525 13016 10897 14021 +FBgn0000039 1775 1433 1594 1531 1653 1776 1280 1695 +FBgn0000042 17647 14551 17832 19160 16484 18013 17594 23033 +FBgn0000043 634 503 571 611 352 337 429 587 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1 0 0 3 +FBgn0264700 0 0 0 0 0 0 0 0 +FBgn0264701 0 1 1 1 0 0 0 2 +FBgn0264702 21 21 15 20 18 21 21 18 +FBgn0264703 10 4 8 7 10 13 7 8 +FBgn0264704 7 8 10 9 5 12 5 3 +FBgn0264705 2 0 1 1 3 1 2 0 +FBgn0264706 0 0 0 0 0 0 0 0 +FBgn0264707 5326 4380 3881 5129 4501 4938 4054 4860 +FBgn0264708 1 0 1 1 2 1 1 0 +FBgn0264709 10 5 7 3 2 3 4 5 +FBgn0264710 0 0 0 0 0 0 0 0 +FBgn0264711 30 19 11 32 47 47 31 27 +FBgn0264712 617 480 488 595 509 533 528 679 +FBgn0264713 1 0 1 1 1 1 2 0 +FBgn0264714 0 0 0 0 0 0 0 0 +FBgn0264715 0 0 0 0 0 0 0 0 +FBgn0264716 0 0 0 0 0 0 0 1 +FBgn0264717 1 0 0 0 0 0 0 2 +FBgn0264718 82 51 31 78 53 69 60 55 +FBgn0264719 0 0 0 0 0 0 0 1 +FBgn0264720 0 0 0 0 0 0 0 0 +FBgn0264721 15 12 17 21 17 16 20 18 +FBgn0264722 3 0 1 1 0 0 0 3 +FBgn0264723 1 0 0 0 1 2 0 0 +FBgn0264724 0 0 0 2 0 0 1 0 +FBgn0264725 0 0 0 0 0 0 1 1 +FBgn0264726 0 1 0 0 0 0 0 2 +FBgn0264727 17 15 9 9 9 8 9 14 diff -r 000000000000 -r a48dde9abecf test-data/vst_transf_htmlout.html --- /dev/null Thu Jan 01 00:00:00 1970 +0000 +++ b/test-data/vst_transf_htmlout.html Wed Jul 17 04:06:12 2013 -0400 @@ -0,0 +1,306 @@ + + + + + + +VST transformation + + + + + + + + + + + + + + + + + + +

VST transformation

+ +
+ +

Loading the R environment

+ +
library("DESeq")
+library(reshape)
+library(ggplot2)
+
+ +

Reading input

+ +
filename = "counttable1.tsv"
+raw_counts = read.csv(filename, sep = "\t", header = T, row.names = 1, stringsAsFactors = F)
+raw_counts_deseqct = newCountDataSet(raw_counts[, c(1, 2)], factor(c("can", 
+    "can")))
+
+ +
+ +

Performing VST transformation

+ +
raw_counts_deseqct = newCountDataSet(raw_counts[, c(1, 2)], factor(c("can", 
+    "can")))
+raw_counts_deseqct = estimateSizeFactors(raw_counts_deseqct)
+raw_counts_deseqctBlind = estimateDispersions(raw_counts_deseqct, method = "blind")
+vsd = varianceStabilizingTransformation(raw_counts_deseqctBlind)
+vst_raw_counts = exprs(vsd)
+
+ +
+ +
write.table(vst_raw_counts, file = "vst_transf_tabout.txt", sep = "\t", quote = F, 
+    col.names = NA, row.names = TRUE)
+head(vst_raw_counts)
+
+ +
##              can_1  can_2
+## FBgn0000003 10.095 10.155
+## FBgn0000008 11.078 11.130
+## FBgn0000014  9.834  9.921
+## FBgn0000015  9.834  9.834
+## FBgn0000017 12.450 12.406
+## FBgn0000018 10.414 10.429
+
+ +
vst_raw_counts_melted = melt(vst_raw_counts)
+ggplot(data = vst_raw_counts_melted, aes(value, colour = X2))
+
+ +
## Error: No layers in plot
+
+ +
+geom_density(alpha = 0.2)
+
+ +
## Error: invalid argument to unary operator
+
+ +
+labs(x = "VST of counts")
+
+ +
## Error: invalid argument to unary operator
+
+ +

+for (sample in colnames(vst_raw_counts)) {
+    hist(vst_raw_counts[, colnames(vst_raw_counts) == sample], main = paste("Sample", 
+        sample), breaks = 80, xlab = "VST counts")
+}
+
+ +

plot of chunk plotting and writing plot of chunk plotting and writing

+ +
+ +
+ +

Logging R environment

+ +
sessionInfo()
+
+ +
## R version 3.0.1 (2013-05-16)
+## Platform: x86_64-pc-linux-gnu (64-bit)
+## 
+## 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=C                 LC_NAME=C                 
+##  [9] LC_ADDRESS=C               LC_TELEPHONE=C            
+## [11] LC_MEASUREMENT=en_US.UTF-8 LC_IDENTIFICATION=C       
+## 
+## attached base packages:
+## [1] parallel  stats     graphics  grDevices utils     datasets  methods  
+## [8] base     
+## 
+## other attached packages:
+##  [1] ggplot2_0.9.3.1    reshape_0.8.4      plyr_1.8          
+##  [4] DESeq_1.12.0       lattice_0.20-15    locfit_1.5-9.1    
+##  [7] Biobase_2.20.1     BiocGenerics_0.6.0 markdown_0.6.1    
+## [10] knitr_1.2         
+## 
+## loaded via a namespace (and not attached):
+##  [1] annotate_1.38.0      AnnotationDbi_1.22.6 colorspace_1.2-2    
+##  [4] DBI_0.2-7            dichromat_2.0-0      digest_0.6.3        
+##  [7] evaluate_0.4.4       formatR_0.8          genefilter_1.42.0   
+## [10] geneplotter_1.38.0   grid_3.0.1           gtable_0.1.2        
+## [13] IRanges_1.18.2       labeling_0.2         MASS_7.3-27         
+## [16] munsell_0.4.2        proto_0.3-10         RColorBrewer_1.0-5  
+## [19] reshape2_1.2.2       RSQLite_0.11.4       scales_0.2.3        
+## [22] splines_3.0.1        stats4_3.0.1         stringr_0.6.2       
+## [25] survival_2.37-4      tools_3.0.1          XML_3.98-1.1        
+## [28] xtable_1.7-1
+
+ +
+ + + + diff -r 000000000000 -r a48dde9abecf test-data/vst_transf_tabout.txt --- /dev/null Thu Jan 01 00:00:00 1970 +0000 +++ b/test-data/vst_transf_tabout.txt Wed Jul 17 04:06:12 2013 -0400 @@ -0,0 +1,15683 @@ + can_1 can_2 +FBgn0000003 10.0947087365926 10.155217055649 +FBgn0000008 11.0778113307378 11.1295635190438 +FBgn0000014 9.83428318137882 9.92126201825076 +FBgn0000015 9.83428318137882 9.83428318137882 +FBgn0000017 12.4499264643294 12.4057034332022 +FBgn0000018 10.413505307596 10.4285708023499 +FBgn0000022 9.83428318137882 9.83428318137882 +FBgn0000024 13.0136340448334 12.9507956467019 +FBgn0000028 10.6787049368095 10.7011923487049 +FBgn0000032 10.6335464215559 10.684563679743 +FBgn0000036 11.1117454091996 11.0186332794786 +FBgn0000037 12.1367362187378 12.1414311086754 +FBgn0000038 13.9975905167321 13.9764048635141 +FBgn0000039 11.6288830936994 11.6194562026446 +FBgn0000042 14.1138264268777 14.1258994844848 +FBgn0000043 10.949357728479 10.9338982360254 +FBgn0000044 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10.0213240287886 10.0286561864236 diff -r 000000000000 -r a48dde9abecf tool_dependencies.xml --- /dev/null Thu Jan 01 00:00:00 1970 +0000 +++ b/tool_dependencies.xml Wed Jul 17 04:06:12 2013 -0400 @@ -0,0 +1,51 @@ + + + + + + + + + + + + + + $INSTALL_DIR + echo "install.packages('reshape')" > $INSTALL_DIR/runme.R + echo "install.packages('knitr')" >> $INSTALL_DIR/runme.R + echo "install.packages('markdown')" >> $INSTALL_DIR/runme.R + echo "quit(save='no')" >> $INSTALL_DIR/runme.R + export PATH=$PATH && export R_HOME=$R_HOME && export R_LIBS=$R_LIBS && + R CMD BATCH $INSTALL_DIR/runme.R > $INSTALL_DIR/install.log + cat $INSTALL_DIR/install.log + + + Installs knitr, reshape and markdown packages for R3.0 + + + + + + + + + $INSTALL_DIR + echo "install.packages('reshape')" > $INSTALL_DIR/runme.R + echo "install.packages('knitr')" >> $INSTALL_DIR/runme.R + echo "install.packages('markdown')" >> $INSTALL_DIR/runme.R + echo "source('http://bioconductor.org/biocLite.R')" >> $INSTALL_DIR/runme.R + echo "installme=c('edgeR','limma','DESeq','DEXSeq','ggplot2','gplots')" >> $INSTALL_DIR/runme.R + echo "biocLite()" >> $INSTALL_DIR/runme.R + echo "biocLite(installme)" >> $INSTALL_DIR/runme.R + echo "quit(save='no')" >> $INSTALL_DIR/runme.R + export PATH=$PATH && export R_HOME=$R_HOME && export R_LIBS=$R_LIBS && + R CMD BATCH $INSTALL_DIR/runme.R > $INSTALL_DIR/install.log + cat $INSTALL_DIR/install.log + + + Installs some basic bioc packages for the edgeR wrapper + It's clunky but this is the only way I could get anything installed into the package_r3 R + + + diff -r 000000000000 -r a48dde9abecf vst_transformation.pl --- /dev/null Thu Jan 01 00:00:00 1970 +0000 +++ b/vst_transformation.pl Wed Jul 17 04:06:12 2013 -0400 @@ -0,0 +1,240 @@ +#!/usr/bin/perl +# vst_transformation.pl +# Joachim Jacob - joachim.jacob@vib.be - 2013 + +use strict; +use File::Basename; +use Log::Log4perl qw(:easy); + +# ---------------------- Prepping Logging -----------------------------# +######################################################################## +# Log levels: $DEBUG $INFO $WARN $ERROR $FATAL +# ConversionPattern: %d %-5p %F{1} [%M] (line %L): %m%n%n +my $log_conf = q/ + log4perl.category = ERROR, Screen + log4perl.appender.Screen.stderr=1 + log4perl.appender.Screen.layout=Log::Log4perl::Layout::PatternLayout + log4perl.appender.Screen.layout.ConversionPattern = %d %-5p %m%n + log4perl.appender.Screen = Log::Log4perl::Appender::Screen +/; + +Log::Log4perl::init( \$log_conf ); +my $logger = get_logger(); + +# ----------------- Getting parameters file ---------------------------# +######################################################################## +my ($parameters) = @ARGV; +my (%para, @repeat_blocks); +open(PARAMETERS,"<$parameters"); +while () { + if (/(\S+)==(.+)$/){ + $para{$1} = $2; + } +} +close(PARAMETERS); + +=Excerpt Config parameters + counttable=$counttable + outtab==$outttab + html_file==$html_file + + ##REPEAT BLOCK + #for $i, $s in enumerate( $replicates ) + replicates_$i==$s.repl_group_name + replicates_$i==$s.rep_columns + #end for + + replicates_$s.repl_group_name==$s.rep_columns + +=cut + +for my $para (keys %para){ + INFO "$para==$para{$para}"; +} + +# --------------------- KNITR SCRIPT ----------------------------------# +######################################################################## +# This script compile the HTML output from the R Markdown script below. +my $htmloutput=$para{'html_file'}; +open(KNITRSCRIPT , ">" , "knitrscript.r"); +print KNITRSCRIPT <>" , "rmarkdownscript.rmd"); + +## Initiating R environment +## --------------------------------------------------------------------- +print RMARKDOWN <){ + INFO $_; +} +close($knitrscript); +close($rmarkdownscript); + + +# -------------------- Assembling command ----------------------------# +######################################################################## +my $command = "R CMD BATCH knitrscript.r"; # to execute + +# --------------------- Executing command ----------------------------# +######################################################################## +run_process($command, "Knitting RMarkdown script"); + +# ------------------- Cleaning up and exiting ------------------------# +######################################################################## +$command = "rm -rf temp.md knitrscript.r knitrscript.r.Rout rmarkdownscript.rmd"; +run_process($command, "Cleaning up"); +exit 0; + +### Subroutines ### +######################################################################## +sub run_process { + my ($command, $name)= @_; + my $logger = get_logger(); + INFO "Launching process\n $command\n"; + system("$command 2>/dev/null") == 0 or die "$name failed\nExit status $?\nCommand: $command"; + if ($? == -1) { + print "failed to execute: $!\n"; + } elsif ($? & 127) { + printf "child died with signal %d, %s coredump\n", ($? & 127), ($? & 128) ? 'with' : 'without'; + } else { + printf "$name executed successfully\n", $? >> 8; + } +} + + + diff -r 000000000000 -r a48dde9abecf vst_transformation.xml --- /dev/null Thu Jan 01 00:00:00 1970 +0000 +++ b/vst_transformation.xml Wed Jul 17 04:06:12 2013 -0400 @@ -0,0 +1,73 @@ + + + using DESeq + + + + package_r3 + r_additions + biocbasics + + + + vst_transformation.pl ${parameters} + + + + + + + + + + + + + + + + + + + + + + + + + + + ## first we pass some galaxy environment variables + counttable==$counttable + header_row==$header_row + header_columns==$header_columns + outtab==$outtab + html_file==$html_file + + ##REPEAT BLOCK + #for $i, $s in enumerate( $replicates ) + replicates_$s.repl_group_name==$s.rep_columns + #end for + + + + + + + + + + + + + + + + + +**Variance stabilizing transformation** + +This tool takes in raw count data per feature and applies a variance stabilizing transformation using DESeq. + +