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GuyTeichman / RNAlysis

Analyze your RNA sequencing data without writing a single line of code
https://guyteichman.github.io/RNAlysis/
MIT License
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DESeq2 fails to execute #21

Closed dw3015 closed 1 year ago

dw3015 commented 1 year ago

Description

Hi there, I am using the stand-alone interface of RNAlysis on MacOS, I have used it with success previously. I am trying to run DESeq2 on the output table generated from kallisto, but it keeps giving me the following error after about 15 seconds of running.

Traceback (most recent call last): File "rnalysis/gui/gui.py", line 4130, in finish_generic_job File "rnalysis/gui/gui_widgets.py", line 375, in run File "rnalysis/filtering.py", line 2864, in differential_expression_deseq2 File "rnalysis/utils/differential_expression.py", line 130, in run_deseq2_analysis File "rnalysis/utils/io.py", line 1416, in run_r_script AssertionError: R script failed to execute (return code 1).

Thanks for any help you can provide!

GuyTeichman commented 1 year ago

Hi dw3015,

Could you please copy the text that appears in the RNAlysis log box after you try to run DESeq2? It should contain a complete report from R/DESeq2 that could help us diagnose the issue further.

Thanks, Guy.

dw3015 commented 1 year ago

Count matrix named 'fed_doubled_kallisto_output_scaled_per_gene.csv' Attaching package: ‘BiocGenerics’ The following objects are masked from ‘package:stats’: IQR, mad, sd, var, xtabs The following objects are masked from ‘package:base’: anyDuplicated, aperm, append, as.data.frame, basename, cbind, colnames, dirname, do.call, duplicated, eval, evalq, Filter, Find, get, grep, grepl, intersect, is.unsorted, lapply, Map, mapply, match, mget, order, paste, pmax, pmax.int, pmin, pmin.int, Position, rank, rbind, Reduce, rownames, sapply, setdiff, sort, table, tapply, union, unique, unsplit, which.max, which.min Attaching package: ‘S4Vectors’ The following object is masked from ‘package:utils’: findMatches The following objects are masked from ‘package:base’: expand.grid, I, unname Attaching package: ‘MatrixGenerics’ The following objects are masked from ‘package:matrixStats’: colAlls, colAnyNAs, colAnys, colAvgsPerRowSet, colCollapse, colCounts, colCummaxs, colCummins, colCumprods, colCumsums, colDiffs, colIQRDiffs, colIQRs, colLogSumExps, colMadDiffs, colMads, colMaxs, colMeans2, colMedians, colMins, colOrderStats, colProds, colQuantiles, colRanges, colRanks, colSdDiffs, colSds, colSums2, colTabulates, colVarDiffs, colVars, colWeightedMads, colWeightedMeans, colWeightedMedians, colWeightedSds, colWeightedVars, rowAlls, rowAnyNAs, rowAnys, rowAvgsPerColSet, rowCollapse, rowCounts, rowCummaxs, rowCummins, rowCumprods, rowCumsums, rowDiffs, rowIQRDiffs, rowIQRs, rowLogSumExps, rowMadDiffs, rowMads, rowMaxs, rowMeans2, rowMedians, rowMins, rowOrderStats, rowProds, rowQuantiles, rowRanges, rowRanks, rowSdDiffs, rowSds, rowSums2, rowTabulates, rowVarDiffs, rowVars, rowWeightedMads, rowWeightedMeans, rowWeightedMedians, rowWeightedSds, rowWeightedVars Welcome to Bioconductor Vignettes contain introductory material; view with 'browseVignettes()'. To cite Bioconductor, see 'citation("Biobase")', and for packages 'citation("pkgname")'. Attaching package: ‘Biobase’ The following object is masked from ‘package:MatrixGenerics’: rowMedians The following objects are masked from ‘package:matrixStats’: anyMissing, rowMedians Loading required package: DESeq2 Loading required package: S4Vectors Loading required package: stats4 Loading required package: BiocGenerics Attaching package: ‘BiocGenerics’ The following objects are masked from ‘package:stats’: IQR, mad, sd, var, xtabs The following objects are masked from ‘package:base’: anyDuplicated, aperm, append, as.data.frame, basename, cbind, colnames, dirname, do.call, duplicated, eval, evalq, Filter, Find, get, grep, grepl, intersect, is.unsorted, lapply, Map, mapply, match, mget, order, paste, pmax, pmax.int, pmin, pmin.int, Position, rank, rbind, Reduce, rownames, sapply, setdiff, sort, table, tapply, union, unique, unsplit, which.max, which.min Attaching package: ‘S4Vectors’ The following object is masked from ‘package:utils’: findMatches The following objects are masked from ‘package:base’: expand.grid, I, unname Loading required package: IRanges Loading required package: GenomicRanges Loading required package: GenomeInfoDb Loading required package: SummarizedExperiment Loading required package: MatrixGenerics Loading required package: matrixStats Attaching package: ‘MatrixGenerics’ The following objects are masked from ‘package:matrixStats’: colAlls, colAnyNAs, colAnys, colAvgsPerRowSet, colCollapse, colCounts, colCummaxs, colCummins, colCumprods, colCumsums, colDiffs, colIQRDiffs, colIQRs, colLogSumExps, colMadDiffs, colMads, colMaxs, colMeans2, colMedians, colMins, colOrderStats, colProds, colQuantiles, colRanges, colRanks, colSdDiffs, colSds, colSums2, colTabulates, colVarDiffs, colVars, colWeightedMads, colWeightedMeans, colWeightedMedians, colWeightedSds, colWeightedVars, rowAlls, rowAnyNAs, rowAnys, rowAvgsPerColSet, rowCollapse, rowCounts, rowCummaxs, rowCummins, rowCumprods, rowCumsums, rowDiffs, rowIQRDiffs, rowIQRs, rowLogSumExps, rowMadDiffs, rowMads, rowMaxs, rowMeans2, rowMedians, rowMins, rowOrderStats, rowProds, rowQuantiles, rowRanges, rowRanks, rowSdDiffs, rowSds, rowSums2, rowTabulates, rowVarDiffs, rowVars, rowWeightedMads, rowWeightedMeans, rowWeightedMedians, rowWeightedSds, rowWeightedVars Loading required package: Biobase Welcome to Bioconductor Vignettes contain introductory material; view with 'browseVignettes()'. To cite Bioconductor, see 'citation("Biobase")', and for packages 'citation("pkgname")'. Attaching package: ‘Biobase’ The following object is masked from ‘package:MatrixGenerics’: rowMedians The following objects are masked from ‘package:matrixStats’: anyMissing, rowMedians converting counts to integer mode Error in contrasts<-(*tmp*, value = contr.funs[1 + isOF[nn]]) : contrasts can be applied only to factors with 2 or more levels Calls: DESeqDataSetFromMatrix -> DESeqDataSet -> -> contrasts<- In addition: Warning message: In DESeqDataSet(se, design = design, ignoreRank) : some variables in design formula are characters, converting to factors Execution halted Attaching package: ‘BiocGenerics’ The following objects are masked from ‘package:stats’: IQR, mad, sd, var, xtabs The following objects are masked from ‘package:base’: anyDuplicated, aperm, append, as.data.frame, basename, cbind, colnames, dirname, do.call, duplicated, eval, evalq, Filter, Find, get, grep, grepl, intersect, is.unsorted, lapply, Map, mapply, match, mget, order, paste, pmax, pmax.int, pmin, pmin.int, Position, rank, rbind, Reduce, rownames, sapply, setdiff, sort, table, tapply, union, unique, unsplit, which.max, which.min Attaching package: ‘S4Vectors’ The following object is masked from ‘package:utils’: findMatches The following objects are masked from ‘package:base’: expand.grid, I, unname Attaching package: ‘MatrixGenerics’ The following objects are masked from ‘package:matrixStats’: colAlls, colAnyNAs, colAnys, colAvgsPerRowSet, colCollapse, colCounts, colCummaxs, colCummins, colCumprods, colCumsums, colDiffs, colIQRDiffs, colIQRs, colLogSumExps, colMadDiffs, colMads, colMaxs, colMeans2, colMedians, colMins, colOrderStats, colProds, colQuantiles, colRanges, colRanks, colSdDiffs, colSds, colSums2, colTabulates, colVarDiffs, colVars, colWeightedMads, colWeightedMeans, colWeightedMedians, colWeightedSds, colWeightedVars, rowAlls, rowAnyNAs, rowAnys, rowAvgsPerColSet, rowCollapse, rowCounts, rowCummaxs, rowCummins, rowCumprods, rowCumsums, rowDiffs, rowIQRDiffs, rowIQRs, rowLogSumExps, rowMadDiffs, rowMads, rowMaxs, rowMeans2, rowMedians, rowMins, rowOrderStats, rowProds, rowQuantiles, rowRanges, rowRanks, rowSdDiffs, rowSds, rowSums2, rowTabulates, rowVarDiffs, rowVars, rowWeightedMads, rowWeightedMeans, rowWeightedMedians, rowWeightedSds, rowWeightedVars Welcome to Bioconductor Vignettes contain introductory material; view with 'browseVignettes()'. To cite Bioconductor, see 'citation("Biobase")', and for packages 'citation("pkgname")'. Attaching package: ‘Biobase’ The following object is masked from ‘package:MatrixGenerics’: rowMedians The following objects are masked from ‘package:matrixStats’: anyMissing, rowMedians Loading required package: DESeq2 Loading required package: S4Vectors Loading required package: stats4 Loading required package: BiocGenerics Attaching package: ‘BiocGenerics’ The following objects are masked from ‘package:stats’: IQR, mad, sd, var, xtabs The following objects are masked from ‘package:base’: anyDuplicated, aperm, append, as.data.frame, basename, cbind, colnames, dirname, do.call, duplicated, eval, evalq, Filter, Find, get, grep, grepl, intersect, is.unsorted, lapply, Map, mapply, match, mget, order, paste, pmax, pmax.int, pmin, pmin.int, Position, rank, rbind, Reduce, rownames, sapply, setdiff, sort, table, tapply, union, unique, unsplit, which.max, which.min Attaching package: ‘S4Vectors’ The following object is masked from ‘package:utils’: findMatches The following objects are masked from ‘package:base’: expand.grid, I, unname Loading required package: IRanges Loading required package: GenomicRanges Loading required package: GenomeInfoDb Loading required package: SummarizedExperiment Loading required package: MatrixGenerics Loading required package: matrixStats Attaching package: ‘MatrixGenerics’ The following objects are masked from ‘package:matrixStats’: colAlls, colAnyNAs, colAnys, colAvgsPerRowSet, colCollapse, colCounts, colCummaxs, colCummins, colCumprods, colCumsums, colDiffs, colIQRDiffs, colIQRs, colLogSumExps, colMadDiffs, colMads, colMaxs, colMeans2, colMedians, colMins, colOrderStats, colProds, colQuantiles, colRanges, colRanks, colSdDiffs, colSds, colSums2, colTabulates, colVarDiffs, colVars, colWeightedMads, colWeightedMeans, colWeightedMedians, colWeightedSds, colWeightedVars, rowAlls, rowAnyNAs, rowAnys, rowAvgsPerColSet, rowCollapse, rowCounts, rowCummaxs, rowCummins, rowCumprods, rowCumsums, rowDiffs, rowIQRDiffs, rowIQRs, rowLogSumExps, rowMadDiffs, rowMads, rowMaxs, rowMeans2, rowMedians, rowMins, rowOrderStats, rowProds, rowQuantiles, rowRanges, rowRanks, rowSdDiffs, rowSds, rowSums2, rowTabulates, rowVarDiffs, rowVars, rowWeightedMads, rowWeightedMeans, rowWeightedMedians, rowWeightedSds, rowWeightedVars Loading required package: Biobase Welcome to Bioconductor Vignettes contain introductory material; view with 'browseVignettes()'. To cite Bioconductor, see 'citation("Biobase")', and for packages 'citation("pkgname")'. Attaching package: ‘Biobase’ The following object is masked from ‘package:MatrixGenerics’: rowMedians The following objects are masked from ‘package:matrixStats’: anyMissing, rowMedians converting counts to integer mode Error in contrasts<-(*tmp*, value = contr.funs[1 + isOF[nn]]) : contrasts can be applied only to factors with 2 or more levels Calls: DESeqDataSetFromMatrix -> DESeqDataSet -> -> contrasts<- In addition: Warning message: In DESeqDataSet(se, design = design, ignoreRank) : some variables in design formula are characters, converting to factors Execution halted

dw3015 commented 1 year ago

I believe I solved it. One group of samples only had a single replicate, and i believe the lack of variance in that group resulted in my error. I was able to pass through the error by removing that group.

GuyTeichman commented 1 year ago

That's good to hear! That's my conclusion as well from reading the DESeq2 log. In the next version of RNAlysis this issue should give out a clearer indication of what went wrong.

Best, Guy.