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  • shikha5
    replied
    Hi @areyes,
    Thanks for your reply. You have correctly pointed out that, the error I was getting was due to the "absence of replicates" in the analysis. But, when I included 2 replicates then it was working fine.

    So, DEXSeq program for estimating dispersion requires atleast 2 repicates per sample.

    Shikha Vashisht

    Leave a comment:


  • areyes
    replied
    Hi @shikha5,

    The error seems to be that a you have no replicates in your experiment.

    As for the dataset with replication, could you add what Mike asked in post 6 and the error that you are getting?

    Alejandro

    Leave a comment:


  • shikha5
    replied
    Hi All,
    Now I am considering the replicates too but getting the same error as mentioned in my last post on this thread.

    Can anybody help?

    Leave a comment:


  • shikha5
    replied
    countFiles = list.files("/dati/data_svashisht/CARIPLO_Project_files/TopHat2_New_alignments_28_11_2014/dexseqCounts", full.names=TRUE)
    flattenedFile = "/personal/svashisht/Cariplo_data_analysis_29-05-14/DEXSeqAnalysis/Homo_sapiens.GRCh37_chr.gff"

    sampleTable = data.frame(row.names = c( "control", "patient"),condition = c("SMA-control", "SMA-patient"),libType = c( "paired-end", "paired-end"))
    library( "DEXSeq" )

    dxd = DEXSeqDataSetFromHTSeq(countFiles,sampleData=sampleTable,design= ~ sample + exon + condition:exon,flattenedfile=flattenedFile )

    colData(dxd)
    DataFrame with 4 rows and 5 columns
    sample condition libType exon sizeFactor
    <factor> <factor> <factor> <factor> <numeric>
    1 control SMA-control paired-end this 1.118398
    2 patient SMA-patient paired-end this 0.894136
    3 control SMA-control paired-end others 1.118398
    4 patient SMA-patient paired-end others 0.894136

    head( rowData(dxd), 3 )
    GRanges object with 3 ranges and 5 metadata columns:
    seqnames ranges strand | featureID
    <Rle> <IRanges> <Rle> | <character>
    ENSG00000000003:E001 X [99883667, 99884983] - | E001
    ENSG00000000003:E002 X [99885756, 99885863] - | E002
    ENSG00000000003:E003 X [99887482, 99887537] - | E003
    groupID exonBaseMean exonBaseVar transcripts
    <character> <numeric> <numeric> <list>
    ENSG00000000003:E001 ENSG00000000003 1054.5 4324.5 ########
    ENSG00000000003:E002 ENSG00000000003 463.0 200.0 ########
    ENSG00000000003:E003 ENSG00000000003 372.0 1352.0 ########
    -------
    seqinfo: 265 sequences from an unspecified genome; no seqlengths
    #Normalization
    dxd <- estimateSizeFactors( dxd )

    #Dispersion Estimation
    dxd <- estimateDispersions (dxd)
    using supplied model matrix
    Error: 1 errors; first error:
    Error in fitBetaWrapper(ySEXP = counts(object), xSEXP = modelMatrix, nfSEXP = normalizationFactors, : in call to fitBeta, the following arguments contain NA: alpha_hatSEXP

    For more information, use bplasterror(). To resume calculation, re-call
    the function and set the argument 'BPRESUME' to TRUE or wrap the
    previous call in bpresume().

    First traceback:
    23: estimateDispersions(dxd)
    22: estimateDispersions(dxd)
    21: .local(object, ...)
    20: bplapply(splitObject, function(x) {
    estimateDispersionsGeneEst(x, maxit = maxit, quiet = quiet,
    modelMatrix = modelMatrix, niter = 10)
    }, BPPARAM = BPPARAM)
    19: bplapply(splitObject, function(x) {
    estimateDispersionsGeneEst(x, maxit = maxit, quiet = quiet,
    modelMatrix = modelMatrix, niter = 10)
    }, BPPARAM = BPPARAM)
    18: mclapply(X = X, FUN = FUN, ..., mc.set.seed = BPPARAM$setSeed,
    mc.silent = !BPPARAM$verbose, mc.cores = bpworkers

    sessionInfo()
    R version 3.1.2 (2014-10-31)
    Platform: x86_64-redhat-linux-gnu (64-bit)

    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] stats4 parallel stats graphics grDevices utils datasets
    [8] methods base

    other attached packages:
    [1] DEXSeq_1.12.1 BiocParallel_1.0.0 DESeq2_1.6.2
    [4] RcppArmadillo_0.4.500.0 Rcpp_0.11.3 GenomicRanges_1.18.3
    [7] GenomeInfoDb_1.2.3 IRanges_2.0.0 S4Vectors_0.4.0
    [10] Biobase_2.26.0 BiocGenerics_0.12.1

    loaded via a namespace (and not attached):
    [1] acepack_1.3-3.3 annotate_1.44.0 AnnotationDbi_1.28.1
    [4] base64enc_0.1-2 BatchJobs_1.5 BBmisc_1.8
    [7] biomaRt_2.22.0 Biostrings_2.34.0 bitops_1.0-6
    [10] brew_1.0-6 checkmate_1.5.0 cluster_1.15.3
    [13] codetools_0.2-9 colorspace_1.2-4 DBI_0.3.1
    [16] digest_0.6.4 fail_1.2 foreach_1.4.2
    [19] foreign_0.8-61 Formula_1.1-2 genefilter_1.48.1
    [22] geneplotter_1.44.0 ggplot2_1.0.0 grid_3.1.2
    [25] gtable_0.1.2 Hmisc_3.14-6 hwriter_1.3.2
    [28] iterators_1.0.7 lattice_0.20-29 latticeExtra_0.6-26
    [31] locfit_1.5-9.1 MASS_7.3-35 munsell_0.4.2
    [34] nnet_7.3-8 plyr_1.8.1 proto_0.3-10
    [37] RColorBrewer_1.0-5 RCurl_1.95-4.3 reshape2_1.4
    [40] rpart_4.1-8 Rsamtools_1.18.2 RSQLite_1.0.0
    [43] scales_0.2.4 sendmailR_1.2-1 splines_3.1.2
    [46] statmod_1.4.20 stringr_0.6.2 survival_2.37-7
    [49] tcltk_3.1.2 tools_3.1.2 XML_3.98-1.1
    [52] xtable_1.7-4 XVector_0.6.0 zlibbioc_1.12.0

    Leave a comment:


  • Michael Love
    replied
    just to repeat, we can't help unless you provide all the information.

    1) show all the code you used, including the dxd object construction.

    2) print out the sessionInfo() and paste it here

    3) (shikha5 has already provided this) as.data.frame(colData(dxd))

    Leave a comment:


  • shikha5
    replied
    I have the same error

    I suspect this error may be due to no replicates.

    as.data.frame(colData(dxd))
    sample condition libType exon sizeFactor
    1 control SMA-control paired-end this 1.118398
    2 patient SMA-patient paired-end this 0.894136
    3 control SMA-control paired-end others 1.118398
    4 patient SMA-patient paired-end others 0.894136

    Can anyone help in fixing this error?

    Thanks in advance

    Shikha
    Last edited by shikha5; 12-16-2014, 09:37 AM.

    Leave a comment:


  • Michael Love
    replied
    hi,

    You need to provide more information for maintainers to help debug.

    Show all the code you used, including the dxd object construction, print out the sessionInfo() and paste it here, and it would be useful to know the column data (at least the structure, if you can't share the actual design information):

    as.data.frame(colData(dxd))

    Leave a comment:


  • shilez
    replied
    I have the same error, and the dxd object is this:
    class: DEXSeqDataSet
    dim: 588047 4
    exptData(0):
    assays(1): counts
    rownames(588047): ENSG00000000003:E001 ENSG00000000003:E002 ...
    ENSG00000261841:E003 ENSG00000261842:E001
    rowData metadata column names(5): featureID groupID exonBaseMean
    exonBaseVar transcripts
    colnames: NULL
    colData names(6): sample X ... exon sizeFactor

    Leave a comment:


  • Michael Love
    replied
    hi,

    Can you show the construction of the dxd object and what the column data looks like?

    Leave a comment:


  • dingdian110
    started a topic DEXseq estimateDispersions error

    DEXseq estimateDispersions error

    Hi there,

    I have been trying to use DEXSeq for alternative splicing analysis. I'm doing analysis following example in the DEXSeq manual. Some errors I got when doing estimateDispersions are as follows:

    > dxd = estimateDispersions( dxd )
    using supplied model matrix
    Error in LastError$store(results = results, is.error = is.error, throw.error = TRUE) :
    Errors occurred during execution. First error message:
    Error in fitBeta(ySEXP = counts(object), xSEXP = modelMatrix, nfSEXP = normalizationFactors, : in call to fitBeta, the following arguments contain NA: alpha_hatSEXP

    For more information, use bplasterror().
    To resume calculation, re-call the function and set the argument 'resume' to TRUE or wrap
    the previous call in bpresume().

    First traceback:
    24: estimateDispersions(dxd)
    23: estimateDispersions(dxd)
    22: .local(object, ...)
    21: bplapply(splitObject, function(x) {
    estimateDispersionsGeneEst(x, maxit = maxit, quiet = quiet,
    modelMatrix = modelMatrix, niter = 10)
    }, BPPARAM = BPPARAM)
    20: bplapply(splitObject, function(x) {
    estimateDispersionsGeneEst(x, maxit = maxit, quiet = quiet,
    modelMatrix = modelMatrix, niter = 10)
    }, BPPARAM = BPPARAM)
    19: bpmapply(FUN, X, MoreArgs = list(...), SIMPLIFY = FALSE, USE.NAMES = FALSE,
    resume = resume, BPPARAM = BPPARAM)
    18: bpma

    Does anyone know what this error mean? and how to fix it?

    thanks a lot!

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