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lcalviell / ORFquant

An R package for Splice-aware quantification of translation using Ribo-seq data
GNU General Public License v3.0
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Error when running run_ORFquant #1

Closed marcasriv closed 4 years ago

marcasriv commented 4 years ago

Hi,

I'm having issues when running run_ORFquant. I successfully run the two previous steps as indicated in the tutorial, and then when I run run_ORFquant with the default parameters except canonical_start_only = FALSE, I get the following error:

Loading annotation and Ribo-seq signal ... Thu Jan 30 13:20:14 2020 Loading annotation and Ribo-seq signal --- Done! Thu Jan 30 13:20:18 2020 Summoning ORFquant with 9330235 P_sites positions over 18323 genomic regions using 30 processor(s) ... Thu Jan 30 13:20:21 2020 Summoning ORFquant --- Done! Thu Jan 30 14:57:15 2020 Exporting ORFquant results ... Thu Jan 30 14:57:15 2020 Error in eval(quote(list(...)), env) : object 'for_ORFquant_file' not found

Thanks a lot!

lcalviell commented 4 years ago

Hi,

Very strange, as apparently the ORF finding and quantification seemed to go through... What is the output of your this run? which files did the script produce? Can you please provide more info about your run? e.g. the exact command you used?

Did you move and/or modify the "for_ORFquant_file" in the middle of the run?

Best, Lorenzo

marcasriv commented 4 years ago

Hi Lorenzo,

Unfortunately I get no output at all from running this command.

The exact command I used is the following:

run_ORFquant("~/genomeAligned.out.bam_for_ORFquant", "~/annot.gtf_Rannot", 30, prefix = for_ORFquant_file, gene_name = NA, gene_id = NA, genomic_region = NA, write_temp_files = T, write_GTF_file = T, write_protein_fasta = T, interactive = T, stn.orf_find.all_starts = T, stn.orf_find.nostarts = F, stn.orf_find.start_sel_cutoff = NA, stn.orf_find.start_sel_cutoff_ave = 0.5,stn.orf_find.cutoff_fr_ave = 0.5, stn.orf_quant.cutoff_cums = NA, stn.orf_quant.cutoff_pct = 2, stn.orf_quant.cutoff_P_sites = NA, unique_reads_only = F, canonical_start_only = FALSE)

I did not modify the for_ORFquant_file in the middle of the run. I've also tried running it with different parameters. I've set write_temp_files = F and interactive = F:

run_ORFquant("~/genomeAligned.out.bam_for_ORFquant", "~/annot.gtf_Rannot", 40, prefix = for_ORFquant_file, gene_name = NA, gene_id = NA, genomic_region = NA, write_temp_files = F, write_GTF_file = T, write_protein_fasta = T, interactive = T, stn.orf_find.all_starts = T, stn.orf_find.nostarts = F, stn.orf_find.start_sel_cutoff = NA, stn.orf_find.start_sel_cutoff_ave = 0.5,stn.orf_find.cutoff_fr_ave = 0.5, stn.orf_quant.cutoff_cums = NA, stn.orf_quant.cutoff_pct = 2, stn.orf_quant.cutoff_P_sites = NA, unique_reads_only = F, canonical_start_only = FALSE)

And I get the following:

Loading annotation and Ribo-seq signal ... Thu Jan 30 17:05:07 2020 Loading annotation and Ribo-seq signal --- Done! Thu Jan 30 17:05:10 2020 Summoning ORFquant with 9330235 P_sites positions over 18323 genomic regions using 40 processor(s) ... Thu Jan 30 17:05:14 2020 Summoning ORFquant --- Done! Thu Jan 30 18:35:40 2020 Exporting ORFquant results ... Thu Jan 30 18:35:40 2020

But still no output file

EDIT

I think I misunderstood the manual 'prefix prefix to use for the output files. Defaults to same as for_ORFquant_file (appends to its filename)'

I ran:

run_ORFquant("~/genomeAligned.out.bam_for_ORFquant", "~/annot.gtf_Rannot", 40, prefix = "~/genomeAligned.out.bam_for_ORFquant", gene_name = NA, gene_id = NA, genomic_region = NA, write_temp_files = F, write_GTF_file = T, write_protein_fasta = T, interactive = T, stn.orf_find.all_starts = T, stn.orf_find.nostarts = F, stn.orf_find.start_sel_cutoff = NA, stn.orf_find.start_sel_cutoff_ave = 0.5,stn.orf_find.cutoff_fr_ave = 0.5, stn.orf_quant.cutoff_cums = NA, stn.orf_quant.cutoff_pct = 2, stn.orf_quant.cutoff_P_sites = NA, unique_reads_only = F, canonical_start_only = FALSE)

This time, I got:

Loading annotation and Ribo-seq signal ... Fri Jan 31 09:50:14 2020 Loading annotation and Ribo-seq signal --- Done! Fri Jan 31 09:50:17 2020 Summoning ORFquant with 9330235 P_sites positions over 18323 genomic regions using 30 processor(s) ... Fri Jan 31 09:50:20 2020 Error in (function (x) : attempt to apply non-function No traceback available Summoning ORFquant --- Done! Fri Jan 31 11:26:52 2020 Exporting ORFquant results ... Fri Jan 31 11:26:52 2020

And a genomeAligned.out.bam_for_ORFquant_tmp_ORFquant_results binary file, but still no GTF file.

This is my systemInfo():

sessionInfo() R version 3.5.2 (2018-12-20) Platform: x86_64-pc-linux-gnu (64-bit) Running under: Ubuntu 18.04.1 LTS

Matrix products: default BLAS: /usr/lib/x86_64-linux-gnu/blas/libblas.so.3.7.1 LAPACK: /usr/lib/x86_64-linux-gnu/lapack/liblapack.so.3.7.1

locale: [1] C

attached base packages: [1] grid parallel stats4 stats graphics grDevices utils [8] datasets methods base

other attached packages: [1] futile.logger_1.4.3 tryCatchLog_1.1.6 [3] BSgenome.Chinese.hamster.PacBio_1.0 ORFquant_0.99.0 [5] rmarkdown_1.13 gridExtra_2.3 [7] knitr_1.22 cowplot_0.9.4 [9] ggplot2_3.1.1 reshape2_1.4.3 [11] GenomicFiles_1.18.0 GenomicAlignments_1.18.1 [13] Rsamtools_1.34.1 SummarizedExperiment_1.12.0 [15] DelayedArray_0.8.0 BiocParallel_1.16.6 [17] matrixStats_0.54.0 multitaper_1.0-14 [19] doMC_1.3.6 iterators_1.0.12 [21] foreach_1.4.7 GenomicFeatures_1.34.8 [23] AnnotationDbi_1.44.0 Biobase_2.42.0 [25] BSgenome_1.50.0 Biostrings_2.50.2 [27] XVector_0.22.0 rtracklayer_1.42.2 [29] GenomicRanges_1.34.0 GenomeInfoDb_1.18.2 [31] IRanges_2.16.0 S4Vectors_0.20.1 [33] BiocGenerics_0.28.0 usethis_1.5.0 [35] devtools_2.0.2

loaded via a namespace (and not attached): [1] bitops_1.0-6 fs_1.2.7 [3] bit64_0.9-7 progress_1.2.0 [5] httr_1.4.0 rprojroot_1.3-2 [7] tools_3.5.2 backports_1.1.4 [9] R6_2.4.0 DBI_1.0.0 [11] lazyeval_0.2.2 colorspace_1.4-1 [13] withr_2.1.2 tidyselect_0.2.5 [15] prettyunits_1.0.2 processx_3.3.0 [17] bit_1.1-14 curl_3.3 [19] compiler_3.5.2 cli_1.1.0 [21] formatR_1.6 desc_1.2.0 [23] scales_1.0.0 callr_3.2.0 [25] stringr_1.4.0 digest_0.6.18 [27] htmltools_0.3.6 pkgconfig_2.0.2 [29] sessioninfo_1.1.1 rlang_0.4.0 [31] RSQLite_2.1.1 dplyr_0.8.0.1 [33] VariantAnnotation_1.28.13 RCurl_1.95-4.12 [35] magrittr_1.5 GenomeInfoDbData_1.2.0 [37] Matrix_1.2-17 Rcpp_1.0.1 [39] munsell_0.5.0 stringi_1.4.3 [41] zlibbioc_1.28.0 pkgbuild_1.0.3 [43] plyr_1.8.4 blob_1.1.1 [45] crayon_1.3.4 lattice_0.20-38 [47] hms_0.4.2 ps_1.3.0 [49] pillar_1.3.1 tcltk_3.5.2 [51] codetools_0.2-16 biomaRt_2.38.0 [53] pkgload_1.0.2 futile.options_1.0.1 [55] XML_3.98-1.19 glue_1.3.1 [57] evaluate_0.13 lambda.r_1.2.3 [59] remotes_2.0.4 testthat_2.2.1 [61] gtable_0.3.0 purrr_0.3.2 [63] assertthat_0.2.1 xfun_0.7 [65] tibble_2.1.1 memoise_1.1.0

Thanks a lot,

Marina

marcasriv commented 4 years ago

Hi Lorenzo, Just a quick update. I've managed to successfully run the script line by line in RStudio.

Best,

Marina

lcalviell commented 4 years ago

Hi Marina, Sometimes I also get the Error in (function (x) : attempt to apply non-function but it's pretty random, and it does not call an actual error state (the script keeps running).

Can you send me the genomeAligned.out.bam_for_ORFquant_tmp_ORFquant_results file for inspection?

Could you get all the output using Rstudio?

Best, Lorenzo