I am trying to use the GAGE pathway analysis tutorial with 3 samples of paired-end RNA-Seq reads (placed in a "tophat_all" directory) that I mapped with TopHat (.bam files have been indexed).
An error occurred at the read count step:
When I tried the same with only one sample I get a different error:
Could someone explain me the error?
Does the error come from my reads (potential unpaired reads, different length...)?
Thanks !
Notes:
(1) I am using the last version of R, Biconductor and its packages. Could it come from the newer versions of the packages that would not fit the tutorial anymore?
(2) Here is the traceback after the error occurred:
An error occurred at the read count step:
Code:
> library(TxDb.Hsapiens.UCSC.hg19.knownGene) > exByGn <- exonsBy(TxDb.Hsapiens.UCSC.hg19.knownGene, "gene") > library(GenomicAlignments) > fls <- list.files("tophat_all/", pattern="bam$", full.names =T) > bamfls <- BamFileList(fls) > flag <- scanBamFlag(isSecondaryAlignment=FALSE, isProperPair=TRUE) > param <- ScanBamParam(flag=flag) > gnCnt <- summarizeOverlaps(exByGn, bamfls, mode="Union", ignore.strand=TRUE, single.end=FALSE, param=param) Error in validObject(.Object) : invalid class "SummarizedExperiment0" object: 'assays' nrow differs from 'mcols' nrow
When I tried the same with only one sample I get a different error:
Code:
> gnCnt <- summarizeOverlaps(exByGn, bamfls, mode="Union", ignore.strand=TRUE, single.end=FALSE, param=param) Error in array(x, c(length(x), 1L), if (!is.null(names(x))) list(names(x), : 'data' must be of a vector type, was 'NULL'
Does the error come from my reads (potential unpaired reads, different length...)?
Thanks !
Notes:
(1) I am using the last version of R, Biconductor and its packages. Could it come from the newer versions of the packages that would not fit the tutorial anymore?
Code:
> sessionInfo() R version 3.2.2 (2015-08-14) Platform: x86_64-pc-linux-gnu (64-bit) locale: [1] C attached base packages: [1] stats4 parallel stats graphics grDevices utils datasets [8] methods base other attached packages: [1] GenomicAlignments_1.6.1 [2] Rsamtools_1.22.0 [3] Biostrings_2.38.0 [4] XVector_0.10.0 [5] SummarizedExperiment_1.0.1 [6] TxDb.Hsapiens.UCSC.hg19.knownGene_3.2.2 [7] GenomicFeatures_1.22.4 [8] AnnotationDbi_1.32.0 [9] Biobase_2.30.0 [10] GenomicRanges_1.22.1 [11] GenomeInfoDb_1.6.1 [12] IRanges_2.4.1 [13] S4Vectors_0.8.2 [14] BiocGenerics_0.16.1 loaded via a namespace (and not attached): [1] zlibbioc_1.16.0 BiocParallel_1.4.0 tools_3.2.2 [4] DBI_0.3.1 lambda.r_1.1.7 futile.logger_1.4.1 [7] rtracklayer_1.30.1 futile.options_1.0.0 bitops_1.0-6 [10] RCurl_1.95-4.7 biomaRt_2.26.0 RSQLite_1.0.0 [13] XML_3.98-1.3
Code:
> traceback() 26: stop(msg, ": ", errors, domain = NA) 25: validObject(.Object) 24: initialize(value, ...) 23: initialize(value, ...) 22: new("SummarizedExperiment0", NAMES = names, elementMetadata = rowData, colData = colData, assays = assays, metadata = as.list(metadata)) 21: new_SummarizedExperiment0(from@assays, names(from@rowRanges), mcols(from@rowRanges), from@colData, from@metadata) 20: asMethod(object) 19: as(object, superClass) 18: slot(x, "assays") 17: is(slot(x, "assays"), "Assays") 16: .valid.SummarizedExperiment0.assays_current(x) 15: valid.func(object) 14: validityMethod(as(object, superClass)) 13: anyStrings(validityMethod(as(object, superClass))) 12: validObject(.Object) 11: initialize(value, ...) 10: initialize(value, ...) 9: new("RangedSummarizedExperiment", rowRanges = rowRanges, colData = colData, assays = assays, elementMetadata = elementMetadata, metadata = as.list(metadata)) 8: .new_RangedSummarizedExperiment(assays, rowRanges, colData, metadata) 7: .local(assays, ...) 6: SummarizedExperiment(assays = SimpleList(counts = counts), rowRanges = features, colData = colData) 5: SummarizedExperiment(assays = SimpleList(counts = counts), rowRanges = features, colData = colData) 4: .dispatchBamFiles(features, reads, mode, match.arg(algorithm), ignore.strand, inter.feature = inter.feature, singleEnd = singleEnd, fragments = fragments, param = param, preprocess.reads = preprocess.reads, ...) 3: .local(features, reads, mode, algorithm, ignore.strand, ...) 2: summarizeOverlaps(exByGn, bamfls, mode = "Union", ignore.strand = TRUE, single.end = FALSE, param = param) 1: summarizeOverlaps(exByGn, bamfls, mode = "Union", ignore.strand = TRUE, single.end = FALSE, param = param)
Comment