Question

easyRNASeq: "Error in IRangesList"

0

Entering edit mode

surjray • 0

@surjray-7556

Last seen 8.2 years ago

United States

Dear all,

I have been running easyRNASeq to count reads for RNA-Seq samples. However I ran into an error with the latest easyRNASeq version. Here is the command I used:

fruMA_male_WCS_male_Unique.count.table <- easyRNASeq( filesDirectory = "./FruMA_Male_WCS_Male/tophat_BAM_files/", organism = "Dmelanogaster", readLength=100L, chr.sizes = "auto", annotationMethod = "gtf", annotationFile = "Drosophila_melanogaster.BDGP5.77.gtf", format = "bam", count = "genes", summarization = "geneModels", filenames = c( "accepted_hits_sorted_FruMA_M_1.bam", "accepted_hits_sorted_FruMA_M_2.bam", "accepted_hits_sorted_FruMA_M_3.bam", "accepted_hits_sorted_WCS_M_1.bam", "accepted_hits_sorted_WCS_M_2.bam", "accepted_hits_sorted_WCS_M_3.bam" ))

This read the annotation file properly, but ran into an issue with computing the gene models:

"

Checking arguments...
Fetching annotations...
Computing gene models...
Error in IRangesList(RL_list > 0) :
all elements in '...' must be IRanges objects"

The traceback() command retrieved the following messages:

5: stop("all elements in '...' must be IRanges objects")
4: IRangesList(RL_list > 0)
3: .geneModelAnnotation(genomicAnnotation(obj), nbCore)
2: easyRNASeq(filesDirectory = "./FruMA_Male_WCS_Male/tophat_BAM_files/",
       organism = "Dmelanogaster", readLength = 100L, chr.sizes = "auto",
       annotationMethod = "gtf", annotationFile = "Drosophila_melanogaster.BDGP5.77.gtf",
       format = "bam", count = "genes", summarization = "geneModels",
       filenames = c("accepted_hits_sorted_FruMA_M_1.bam", "accepted_hits_sorted_FruMA_M_2.bam",
           "accepted_hits_sorted_FruMA_M_3.bam", "accepted_hits_sorted_WCS_M_1.bam",
           "accepted_hits_sorted_WCS_M_2.bam", "accepted_hits_sorted_WCS_M_3.bam"))
1: easyRNASeq(filesDirectory = "./FruMA_Male_WCS_Male/tophat_BAM_files/",
       organism = "Dmelanogaster", readLength = 100L, chr.sizes = "auto",
       annotationMethod = "gtf", annotationFile = "Drosophila_melanogaster.BDGP5.77.gtf",
       format = "bam", count = "genes", summarization = "geneModels",
       filenames = c("accepted_hits_sorted_FruMA_M_1.bam", "accepted_hits_sorted_FruMA_M_2.bam",
           "accepted_hits_sorted_FruMA_M_3.bam", "accepted_hits_sorted_WCS_M_1.bam",
           "accepted_hits_sorted_WCS_M_2.bam", "accepted_hits_sorted_WCS_M_3.bam"))"

This very easyRNASeq command, though, runs on the older versions. Can you please help.

Thanking you.

Here is my R session info.

R version 3.2.0 (2015-04-16)
Platform: x86_64-pc-linux-gnu (64-bit)
Running under: Ubuntu 14.04.2 LTS

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] easyRNASeq_2.4.0 locfit_1.5-9.1

loaded via a namespace (and not attached):
[1] genefilter_1.50.0         reshape2_1.4.1
[3] splines_3.2.0             lattice_0.20-31
[5] colorspace_1.2-6          stats4_3.2.0
[7] survival_2.38-1           XML_3.98-1.2
[9] foreign_0.8-63            DBI_0.3.1
[11] BiocParallel_1.2.2        BiocGenerics_0.14.0
[13] RColorBrewer_1.1-2        lambda.r_1.1.7
[15] plyr_1.8.2                stringr_1.0.0
[17] zlibbioc_1.14.0           Biostrings_2.36.1
[19] munsell_0.4.2             gtable_0.1.2
[21] futile.logger_1.4.1       hwriter_1.3.2
[23] DESeq2_1.8.1              latticeExtra_0.6-26
[25] Biobase_2.28.0            RcppArmadillo_0.5.200.1.0
[27] biomaRt_2.24.0            geneplotter_1.46.0
[29] IRanges_2.2.3             GenomeInfoDb_1.4.0
[31] parallel_3.2.0            AnnotationDbi_1.30.1
[33] proto_0.3-10              Rcpp_0.11.6
[35] edgeR_3.10.2              acepack_1.3-3.3
[37] xtable_1.7-4              scales_0.2.4
[39] limma_3.24.8              S4Vectors_0.6.0
[41] Hmisc_3.16-0              annotate_1.46.0
[43] XVector_0.8.0             ShortRead_1.26.0
[45] Rsamtools_1.20.4          gridExtra_0.9.1
[47] ggplot2_1.0.1             digest_0.6.8
[49] stringi_0.4-1             DESeq_1.20.0
[51] GenomicRanges_1.20.4      grid_3.2.0
[53] tools_3.2.0               bitops_1.0-6
[55] magrittr_1.5              RCurl_1.95-4.6
[57] LSD_3.0                   RSQLite_1.0.0
[59] Formula_1.2-1             cluster_2.0.1
[61] futile.options_1.0.0      MASS_7.3-40
[63] rpart_4.1-9               intervals_0.15.0
[65] genomeIntervals_1.24.0    GenomicAlignments_1.4.1
[67] nnet_7.3-9

easyRNASeq • 1.7k views

ADD COMMENT • link 8.9 years ago surjray • 0

score 0 · Answer 1 · 2015-06-10

Hej surjray! Did it work with the exact same annotation file: Drosophila_melanogaster.BDGP5.77.gtf? The error seem to occur when computing the synthetic transcripts. I'll take a closer look asap. Cheers, Nico --------------------------------------------------------------- Nicolas Delhomme, PhD The Street Lab Department of Plant Physiology Umeå Plant Science Center Tel: +46 90 786 5478 Email: nicolas.delhomme@umu.se SLU - Umeå universitet Umeå S-901 87 Sweden --------------------------------------------------------------- > On 09 Jun 2015, at 22:56, surjray [bioc] <noreply@bioconductor.org> wrote: > > Activity on a post you are following on support.bioconductor.org > User surjray wrote Question: easyRNASeq: "Error in IRangesList": > > > Dear all, > > I have been running easyRNASeq to count reads for RNA-Seq samples. However I ran into an error with the latest easyRNASeq version. Here is the command I used: > > fruMA_male_WCS_male_Unique.count.table <- easyRNASeq( filesDirectory = "./FruMA_Male_WCS_Male/tophat_BAM_files/", organism = "Dmelanogaster", readLength=100L, chr.sizes = "auto", annotationMethod = "gtf", annotationFile = "Drosophila_melanogaster.BDGP5.77.gtf", format = "bam", count = "genes", summarization = "geneModels", filenames = c( "accepted_hits_sorted_FruMA_M_1.bam", "accepted_hits_sorted_FruMA_M_2.bam", "accepted_hits_sorted_FruMA_M_3.bam", "accepted_hits_sorted_WCS_M_1.bam", "accepted_hits_sorted_WCS_M_2.bam", "accepted_hits_sorted_WCS_M_3.bam" )) > > This read the annotation file properly, but ran into an issue with computing the gene models: > > " > > Checking arguments... > Fetching annotations... > Computing gene models... > Error in IRangesList(RL_list > 0) : > all elements in '...' must be IRanges objects" > > The traceback() command retrieved the following messages: > > 5: stop("all elements in '...' must be IRanges objects") > 4: IRangesList(RL_list > 0) > 3: .geneModelAnnotation(genomicAnnotation(obj), nbCore) > 2: easyRNASeq(filesDirectory = "./FruMA_Male_WCS_Male/tophat_BAM_files/", > organism = "Dmelanogaster", readLength = 100L, chr.sizes = "auto", > annotationMethod = "gtf", annotationFile = "Drosophila_melanogaster.BDGP5.77.gtf", > format = "bam", count = "genes", summarization = "geneModels", > filenames = c("accepted_hits_sorted_FruMA_M_1.bam", "accepted_hits_sorted_FruMA_M_2.bam", > "accepted_hits_sorted_FruMA_M_3.bam", "accepted_hits_sorted_WCS_M_1.bam", > "accepted_hits_sorted_WCS_M_2.bam", "accepted_hits_sorted_WCS_M_3.bam")) > 1: easyRNASeq(filesDirectory = "./FruMA_Male_WCS_Male/tophat_BAM_files/", > organism = "Dmelanogaster", readLength = 100L, chr.sizes = "auto", > annotationMethod = "gtf", annotationFile = "Drosophila_melanogaster.BDGP5.77.gtf", > format = "bam", count = "genes", summarization = "geneModels", > filenames = c("accepted_hits_sorted_FruMA_M_1.bam", "accepted_hits_sorted_FruMA_M_2.bam", > "accepted_hits_sorted_FruMA_M_3.bam", "accepted_hits_sorted_WCS_M_1.bam", > "accepted_hits_sorted_WCS_M_2.bam", "accepted_hits_sorted_WCS_M_3.bam"))" > > This very easyRNASeq command, though, runs on the older versions. Can you please help. > > Thanking you. > > > Here is my R session info. > > R version 3.2.0 (2015-04-16) > Platform: x86_64-pc-linux-gnu (64-bit) > Running under: Ubuntu 14.04.2 LTS > > 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] easyRNASeq_2.4.0 locfit_1.5-9.1 > > loaded via a namespace (and not attached): > [1] genefilter_1.50.0 reshape2_1.4.1 > [3] splines_3.2.0 lattice_0.20-31 > [5] colorspace_1.2-6 stats4_3.2.0 > [7] survival_2.38-1 XML_3.98-1.2 > [9] foreign_0.8-63 DBI_0.3.1 > [11] BiocParallel_1.2.2 BiocGenerics_0.14.0 > [13] RColorBrewer_1.1-2 lambda.r_1.1.7 > [15] plyr_1.8.2 stringr_1.0.0 > [17] zlibbioc_1.14.0 Biostrings_2.36.1 > [19] munsell_0.4.2 gtable_0.1.2 > [21] futile.logger_1.4.1 hwriter_1.3.2 > [23] DESeq2_1.8.1 latticeExtra_0.6-26 > [25] Biobase_2.28.0 RcppArmadillo_0.5.200.1.0 > [27] biomaRt_2.24.0 geneplotter_1.46.0 > [29] IRanges_2.2.3 GenomeInfoDb_1.4.0 > [31] parallel_3.2.0 AnnotationDbi_1.30.1 > [33] proto_0.3-10 Rcpp_0.11.6 > [35] edgeR_3.10.2 acepack_1.3-3.3 > [37] xtable_1.7-4 scales_0.2.4 > [39] limma_3.24.8 S4Vectors_0.6.0 > [41] Hmisc_3.16-0 annotate_1.46.0 > [43] XVector_0.8.0 ShortRead_1.26.0 > [45] Rsamtools_1.20.4 gridExtra_0.9.1 > [47] ggplot2_1.0.1 digest_0.6.8 > [49] stringi_0.4-1 DESeq_1.20.0 > [51] GenomicRanges_1.20.4 grid_3.2.0 > [53] tools_3.2.0 bitops_1.0-6 > [55] magrittr_1.5 RCurl_1.95-4.6 > [57] LSD_3.0 RSQLite_1.0.0 > [59] Formula_1.2-1 cluster_2.0.1 > [61] futile.options_1.0.0 MASS_7.3-40 > [63] rpart_4.1-9 intervals_0.15.0 > [65] genomeIntervals_1.24.0 GenomicAlignments_1.4.1 > [67] nnet_7.3-9 > > > You may reply via email or visit easyRNASeq: "Error in IRangesList" >

score 0 · Answer 2 · 2015-06-10

0

Entering edit mode

surjray • 0

@surjray-7556

Last seen 8.2 years ago

United States

Dear Nico,

I used the exact same annotation file with older versions of easyRNASeq and they worked fine. The latest version seems to be running into this error.

Thanking you.

ADD COMMENT • link 8.9 years ago surjray • 0

0

Entering edit mode

Hej Surjray! Sorry for the long delay in answering. Just to let you know that I'm looking into it now. Cheers, Nico --------------------------------------------------------------- Nicolas Delhomme, PhD The Street Lab Department of Plant Physiology Umeå Plant Science Center Tel: +46 90 786 5478 Email: nicolas.delhomme@umu.se SLU - Umeå universitet Umeå S-901 87 Sweden --------------------------------------------------------------- > On 10 Jun 2015, at 17:06, surjray [bioc] <noreply@bioconductor.org> wrote: > > Activity on a post you are following on support.bioconductor.org > User surjray wrote Answer: easyRNASeq: "Error in IRangesList": > > > Dear Nico, > > I used the exact same annotation file with older versions of easyRNASeq and they worked fine. The latest version seems to be running into this error. > > Thanking you. > > > You may reply via email or visit A: easyRNASeq: "Error in IRangesList" >

ADD REPLY • link 8.8 years ago Nicolas Delhomme ▴ 320

0

Entering edit mode

Hej Surjray! The method you have been using is deprecated at many levels. I'll try to fix it (it should be defunct by now really...) but meanwhile you should try to move your code to the latest implementation. For that, you should look at the section 7.1 in the vignette to transform your annotation into a "flattened" annotation, one that does avoid most of the double-counting issue. See also these guidelines for more info: http://www.epigenesys.eu/en/protocols/bio-informatics/1283-guidelines-for-rna-seq-data-analysis. Next you should use the simpleRNASeq function as follows: ## Create a list of bamfiles bamFiles <- getBamFileList(filenames= dir(system.file(package="RnaSeqTutorial","extdata"), pattern="*.bam$", full.names=TRUE)) ## create a set of parameters - most of them are actually determined at runtime anyway (e.g. read length, paired-end, etc.) param <- RnaSeqParam( annotParam=AnnotParam(datasource="~/Downloads/Drosophila_melanogaster.BDGP5.77.gtf.gz", type="gtf")) ## execute to get a summarized experiment object sexp <- simpleRNASeq(bamFiles=bamFiles,param=param,verbose=TRUE) HTH, Meanwhile, I'll try to fix the deprecated way one last time; after which it is going to be defunct. I've already fixed your original issue, but I need to ensure that the obtained results are correct. Cheers, Nico --------------------------------------------------------------- Nicolas Delhomme, PhD The Street Lab Department of Plant Physiology Umeå Plant Science Center Tel: +46 90 786 5478 Email: nicolas.delhomme@umu.se SLU - Umeå universitet Umeå S-901 87 Sweden --------------------------------------------------------------- > On 10 Jun 2015, at 17:06, surjray [bioc] <noreply@bioconductor.org> wrote: > > Activity on a post you are following on support.bioconductor.org > > User surjray<https: support.bioconductor.org="" u="" 7556=""/> wrote Answer: easyRNASeq: "Error in IRangesList"<https: support.bioconductor.org="" p="" 68590="" #68620="">: > > Dear Nico, > > I used the exact same annotation file with older versions of easyRNASeq and they worked fine. The latest version seems to be running into this error. > > Thanking you. > > ________________________________ > > You may reply via email or visit A: easyRNASeq: "Error in IRangesList"

ADD REPLY • link 8.8 years ago Nicolas Delhomme ▴ 320

0

Entering edit mode

Hej Surjray! In our public Git repository, I have added a folder: https://microasp.upsc.se/root/upscb-public/tree/master/tutorial/easyRNASeq that contains an example of how I would create the synthetic transcript from the FlyBase gtf file. To do so, just download and look at the html file. You can actually reproduce it using the companion R file. The function that creates the synthetic transcripts is available from the same Git repos; there: https://microasp.upsc.se/root/upscb-public/tree/master/src/R HTH, Cheers, Nico --------------------------------------------------------------- Nicolas Delhomme, PhD The Street Lab Department of Plant Physiology Umeå Plant Science Center Tel: +46 90 786 5478 Email: nicolas.delhomme@umu.se SLU - Umeå universitet Umeå S-901 87 Sweden --------------------------------------------------------------- > On 25 Jun 2015, at 10:01, Nicolas Delhomme [bioc] <noreply@bioconductor.org> wrote: > > Activity on a post you are following on support.bioconductor.org > User Nicolas Delhomme wrote Comment: easyRNASeq: "Error in IRangesList": > > > Hej Surjray! The method you have been using is deprecated at many levels. I'll try to fix it (it should be defunct by now really...) but meanwhile you should try to move your code to the latest implementation. For that, you should look at the section 7.1 in the vignette to transform your annotation into a "flattened" annotation, one that does avoid most of the double-counting issue. See also these guidelines for more info: http://www.epigenesys.eu/en/protocols/bio-informatics/1283-guidelines-for-rna-seq-data-analysis. Next you should use the simpleRNASeq function as follows: ## Create a list of bamfiles bamFiles <- getBamFileList(filenames= dir(system.file(package="RnaSeqTutorial","extdata"), pattern="*.bam$", full.names=TRUE)) ## create a set of parameters - most of them are actually determined at runtime anyway (e.g. read length, paired-end, etc.) param <- RnaSeqParam( annotParam=AnnotParam(datasource="~/Downloads/Drosophila_melanogaster.BDGP5.77.gtf.gz", type="gtf")) ## execute to get a summarized experiment object sexp <- simpleRNASeq(bamFiles=bamFiles,param=param,verbose=TRUE) HTH, Meanwhile, I'll try to fix the deprecated way one last time; after which it is going to be defunct. I've already fixed your original issue, but I need to ensure that the obtained results are correct. Cheers, Nico --------------------------------------------------------------- Nicolas Delhomme, PhD The Street Lab Department of Plant Physiology Umeå Plant Science Center Tel: +46 90 786 5478 Email: nicolas.delhomme@umu.se SLU - Umeå universitet Umeå S-901 87 Sweden --------------------------------------------------------------- > On 10 Jun 2015, at 17:06, surjray [bioc] <noreply@bioconductor.org> wrote: > > Activity on a post you are following on support.bioconductor.org > > User surjray<https: support.bioconductor.org="" u="" 7556=""/> wrote Answer: easyRNASeq: "Error in IRangesList"<https: support.bioconductor.org="" p="" 68590="" #68620="">: > > Dear Nico, > > I used the exact same annotation file with older versions of easyRNASeq and they worked fine. The latest version seems to be running into this error. > > Thanking you. > > ________________________________ > > You may reply via email or visit A: easyRNASeq: "Error in IRangesList" > > Post tags: easyRNASeq > > You may reply via email or visit C: easyRNASeq: "Error in IRangesList" >

ADD REPLY • link 8.8 years ago Nicolas Delhomme ▴ 320

0

Entering edit mode

Sorry, I forgot to say that this will only work with easyRNASeq version 2.4.5, which will be available in about 2 days from Bioc (or immediately if you get the source code from the Bioc SVN repos). Cheers, Nico --------------------------------------------------------------- Nicolas Delhomme, PhD The Street Lab Department of Plant Physiology Umeå Plant Science Center Tel: +46 90 786 5478 Email: nicolas.delhomme@umu.se SLU - Umeå universitet Umeå S-901 87 Sweden --------------------------------------------------------------- > On 07 Jul 2015, at 10:55, Nicolas Delhomme <nicolas.delhomme@umu.se> wrote: > > Hej Surjray! > > In our public Git repository, I have added a folder: https://microasp.upsc.se/root/upscb-public/tree/master/tutorial/easyRNASeq that contains an example of how I would create the synthetic transcript from the FlyBase gtf file. To do so, just download and look at the html file. You can actually reproduce it using the companion R file. The function that creates the synthetic transcripts is available from the same Git repos; there: https://microasp.upsc.se/root/upscb-public/tree/master/src/R > > HTH, > > Cheers, > > Nico > > --------------------------------------------------------------- > Nicolas Delhomme, PhD > > The Street Lab > Department of Plant Physiology > Umeå Plant Science Center > > Tel: +46 90 786 5478 > Email: nicolas.delhomme@umu.se > SLU - Umeå universitet > Umeå S-901 87 Sweden > --------------------------------------------------------------- > >> On 25 Jun 2015, at 10:01, Nicolas Delhomme [bioc] <noreply@bioconductor.org> wrote: >> >> Activity on a post you are following on support.bioconductor.org >> User Nicolas Delhomme wrote Comment: easyRNASeq: "Error in IRangesList": >> >> >> Hej Surjray! The method you have been using is deprecated at many levels. I'll try to fix it (it should be defunct by now really...) but meanwhile you should try to move your code to the latest implementation. For that, you should look at the section 7.1 in the vignette to transform your annotation into a "flattened" annotation, one that does avoid most of the double-counting issue. See also these guidelines for more info: http://www.epigenesys.eu/en/protocols/bio-informatics/1283-guidelines-for-rna-seq-data-analysis. Next you should use the simpleRNASeq function as follows: ## Create a list of bamfiles bamFiles <- getBamFileList(filenames= dir(system.file(package="RnaSeqTutorial","extdata"), pattern="*.bam$", full.names=TRUE)) ## create a set of parameters - most of them are actually determined at runtime anyway (e.g. read length, paired-end, etc.) param <- RnaSeqParam( annotParam=AnnotParam(datasource="~/Downloads/Drosophila_melanogaster.BDGP5.77.gtf.gz", type="gtf")) ## execute to get a summarized experiment object sexp <- simpleRNASeq(bamFiles=bamFiles,param=param,verbose=TRUE) HTH, Meanwhile, I'll try to fix the deprecated way one last time; after which it is going to be defunct. I've already fixed your original issue, but I need to ensure that the obtained results are correct. Cheers, Nico --------------------------------------------------------------- Nicolas Delhomme, PhD The Street Lab Department of Plant Physiology Umeå Plant Science Center Tel: +46 90 786 5478 Email: nicolas.delhomme@umu.se SLU - Umeå universitet Umeå S-901 87 Sweden --------------------------------------------------------------- > On 10 Jun 2015, at 17:06, surjray [bioc] <noreply@bioconductor.org> wrote: > > Activity on a post you are following on support.bioconductor.org > > User surjray<https: support.bioconductor.org="" u="" 7556=""/> wrote Answer: easyRNASeq: "Error in IRangesList"<https: support.bioconductor.org="" p="" 68590="" #68620="">: > > Dear Nico, > > I used the exact same annotation file with older versions of easyRNASeq and they worked fine. The latest version seems to be running into this error. > > Thanking you. > > ________________________________ > > You may reply via email or visit A: easyRNASeq: "Error in IRangesList" >> >> Post tags: easyRNASeq >> >> You may reply via email or visit C: easyRNASeq: "Error in IRangesList" >> >

ADD REPLY • link 8.8 years ago Nicolas Delhomme ▴ 320