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sakura.nussbaum
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@sakuranussbaum-10160
Last seen 8.6 years ago
Hello,
I am a new user of DEXSeq and I would need some help concerning an error message I get and I cannot solve.
My sample table looks like this:
> phenodata
samplename condition
bu1 bulge_1 bulge
bu2 bulge_2 bulge
bu3 bulge_3 bulge
le1 lef1_1 lef1
le4 lef1_4 lef1
le5 lef1_5 lef1
le6 lef1_6 lef1
I am trying to create an ExonCountSet object but I get an error message:
> library(DEXSeq)
> ec = HTSeqCounts(countfiles=c("bulge_1.txt","bulge_2.txt","bulge_3.txt","lef_1.txt","lef_4.txt","lef_5.txt","lef_6.txt"), design=phenodata)
Error: could not find function "HTSeqCounts"
Even if the DEXSeq package seems to be loaded as seen in the session Info:
R version 3.2.3 (2015-12-10) Platform: x86_64-apple-darwin13.4.0 (64-bit) Running under: OS X 10.11.4 (El Capitan) locale: [1] C attached base packages: [1] stats4 parallel stats graphics grDevices utils datasets methods base other attached packages: [1] BiocInstaller_1.20.1 DEXSeq_1.16.10 DESeq2_1.10.1 [4] RcppArmadillo_0.6.700.3.0 Rcpp_0.12.4 SummarizedExperiment_1.0.2 [7] GenomicRanges_1.22.4 GenomeInfoDb_1.6.3 IRanges_2.4.8 [10] S4Vectors_0.8.11 Biobase_2.30.0 BiocGenerics_0.16.1 [13] BiocParallel_1.4.3 loaded via a namespace (and not attached): [1] genefilter_1.52.1 statmod_1.4.24 locfit_1.5-9.1 splines_3.2.3 [5] lattice_0.20-33 colorspace_1.2-6 survival_2.39-2 XML_3.98-1.4 [9] foreign_0.8-66 DBI_0.3.1 RColorBrewer_1.1-2 lambda.r_1.1.7 [13] plyr_1.8.3 stringr_1.0.0 zlibbioc_1.16.0 Biostrings_2.38.4 [17] munsell_0.4.3 gtable_0.2.0 futile.logger_1.4.1 hwriter_1.3.2 [21] latticeExtra_0.6-28 geneplotter_1.48.0 biomaRt_2.26.1 AnnotationDbi_1.32.3 [25] acepack_1.3-3.3 xtable_1.8-2 scales_0.4.0 Hmisc_3.17-3 [29] annotate_1.48.0 XVector_0.10.0 Rsamtools_1.22.0 gridExtra_2.2.1 [33] ggplot2_2.1.0 stringi_1.0-1 grid_3.2.3 tools_3.2.3 [37] bitops_1.0-6 magrittr_1.5 RCurl_1.95-4.8 RSQLite_1.0.0 [41] Formula_1.2-1 cluster_2.0.4 futile.options_1.0.0 Matrix_1.2-5 [45] rsconnect_0.4.2.1 rpart_4.1-10 nnet_7.3-12
Does anyone know why I have this error message?
Thanks in advance for your help,
Sakura
I realised that I had an error, I was supposed to type "read.HTSeqCounts()" instead of just "HTSeqCounts()", but id did not solve my problem, I still get:
Error: could not find function "read.HTSeqCounts"
I am trying to follow the pipeline from this source, and it seems that this function is in the package...
https://books.google.co.jp/books?id=f4tqBAAAQBAJ&pg=PA184&lpg=PA184&dq=Reading+all+the+sample-specific+count+tables+is+done+by+the+function+read.HTSeqCounts()+from+the+package+DEXSeq&source=bl&ots=7lMhp7fVjQ&sig=r6dEc-4slTppnDK8ODVD4RMzJHg&hl=fr&sa=X&ved=0ahUKEwii3vuu46HMAhWGtpQKHaoRDtYQ6AEIHjAA#v=onepage&q=Reading%20all%20the%20sample-specific%20count%20tables%20is%20done%20by%20the%20function%20read.HTSeqCounts()%20from%20the%20package%20DEXSeq&f=false
I think the function has been renamed as
DEXSeqDataSetFromHTSeq(), and the chapter that you are using is (already, unfortunately) out-of-date (I couldn't find via online search the version of DEXSeq used in the chapter -- nosessionInfo()). Some archaeology shows that read.HTSeqCount was 'deprecated' on 2014-03-21, in version 1.9.7 of the package, so more than two years ago; the current version of the package is 1.16.10. The message accompanying the change isso there may be additional differences between the text of the book that you are reading and the package. The good news is that there is an extensive current vignette (also
browseVignettes(package="DEXSeq")) in the package itself.And this is written in that vignette about HTSeq
The initial steps of a DEXSeq analysis, described in the following two sections, is typically done outside R, by using two provided Python scripts
Thank you all for your answers. I changed the method and I am now following this pipeline as you suggested. However, I also have some problems.
When I call the DEXSeqDataSetFromHTSeq, I obtain the following error:
Here is what countFiles and sampleTable look like:
> countFiles [1] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/basal_1.txt" [2] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/basal_2.txt" [3] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/basal_3.txt" [4] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/bulge_1.txt" [5] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/bulge_2.txt" [6] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/bulge_3.txt" [7] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/cdh3_1.txt" [8] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/cdh3_2.txt" [9] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/cdh3_3.txt" [10] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/cdh3_4.txt" [11] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/gli_1.txt" [12] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/gli_2.txt" [13] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/gli_4.txt" [14] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/lef_1.txt" [15] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/lef_4.txt" [16] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/lef_5.txt" [17] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/lef_6.txt" [18] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/lgr6_10_1.txt" [19] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/lgr6_10_2.txt" [20] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/lgr6_40_1.txt" [21] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/lgr6_40_2.txt" [22] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/lgr6_40_3.txt" [23] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/pdgfra_10_1.txt" [24] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/pdgfra_10_2.txt" [25] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/pdgfra_10_3.txt" [26] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/pdgfra_40_1.txt" [27] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/pdgfra_40_2.txt" [28] "/Volumes/TOSHIBAEXT/3.mapping-hisat/Counts/pdgfra_40_3.txt" > sampleTable condition libType basal1 basal single-end basal2 basal single-end basal3 basal single-end bulge1 bulge single-end bulge2 bulge single-end bulge3 bulge single-end cdh31 cdh3 single-end cdh32 cdh3 single-end cdh33 cdh3 single-end cdh34 cdh3 single-end gli11 gli1 single-end gli12 gli1 single-end gli14 gli1 single-end lef11 lef1 single-end lef14 lef1 single-end lef15 lef1 single-end lef16 lef1 single-end lgr6101 lgr6 single-end lgr6102 lgr6 single-end lgr6401 lgr6 single-end lgr6402 lgr6 single-end lgr6403 lgr6 single-end pdgfra101 pdgfra single-end pdgfra102 pdgfra single-end pdgfra103 pdgfra single-end pdgfra401 pdgfra single-end pdgfra402 pdgfra single-end pdgfra403 pdgfra single-endI guess this is correct. However, when looking at the count files, I have the impression that the .txt look weird. Here is the first one (sample: basal_1) as an example. This is all that is contained in the file:
_ambiguous 0
_ambiguous_readpair_position 0
_empty 9651846
_lowaqual 846199
_notaligned 421258
I created this file with this command (and successfully installed pysam before):
Is this output normal with dexseq_count.py? If not, does anyone know what could be the problem? I tried with different options, but as my data are not paired-end and strand specific, I should use is like this.