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3.7 years ago
Aspire
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I retrieve DiffBind results as a DESeq2 object, and then look how many differentially bound regions are within it. When doing this, I get a different number of DB regions compared to the original DiffBind object.
library(DESeq2)
library(DiffBind)
data(tamoxifen_counts)
tamoxifen <- dba.contrast(tamoxifen, design="~Tissue")
tamoxifen <- dba.contrast(tamoxifen, group1=tamoxifen$masks$MCF7, group2=tamoxifen$masks$BT474, name1="MCF7", name2="BT474")
tamoxifen <- dba.analyze(tamoxifen)
dds <- dba.analyze(tamoxifen,bRetrieveAnalysis = DBA_DESEQ2)
res <- results(dds, independentFiltering=F, contrast=c("Tissue","MCF7","BT474"))
dba.show(tamoxifen,bContrasts = T,th=0.1)
summary(res)
sessionInfo( )
Output :
Group Samples Group2 Samples2 DB.DESeq2
1 MCF7 5 BT474 2 954
out of 2845 with nonzero total read count
adjusted p-value < 0.1
LFC > 0 (up) : 738, 26%
LFC < 0 (down) : 437, 15%
outliers [1] : 0, 0%
low counts [2] : 0, 0%
(mean count < 0)
[1] see 'cooksCutoff' argument of ?results
[2] see 'independentFiltering' argument of ?results
R version 4.0.4 (2021-02-15)
Platform: x86_64-pc-linux-gnu (64-bit)
Running under: Ubuntu 20.04.2 LTS
Matrix products: default
BLAS: /usr/lib/x86_64-linux-gnu/blas/libblas.so.3.9.0
LAPACK: /usr/lib/x86_64-linux-gnu/lapack/liblapack.so.3.9.0
locale:
[1] LC_CTYPE=en_IL.UTF-8 LC_NUMERIC=C LC_TIME=en_IL.UTF-8
[4] LC_COLLATE=en_IL.UTF-8 LC_MONETARY=en_IL.UTF-8 LC_MESSAGES=en_IL.UTF-8
[7] LC_PAPER=en_IL.UTF-8 LC_NAME=C LC_ADDRESS=C
[10] LC_TELEPHONE=C LC_MEASUREMENT=en_IL.UTF-8 LC_IDENTIFICATION=C
attached base packages:
[1] parallel stats4 grid stats graphics grDevices utils datasets methods
[10] base
other attached packages:
[1] DiffBind_3.0.13 rgl_0.105.13 limma_3.46.0
[4] DESeq2_1.30.1 SummarizedExperiment_1.20.0 Biobase_2.50.0
[7] MatrixGenerics_1.2.1 matrixStats_0.58.0 GenomicRanges_1.42.0
[10] GenomeInfoDb_1.26.2 IRanges_2.24.1 S4Vectors_0.28.1
[13] BiocGenerics_0.36.0 RColorBrewer_1.1-2 pheatmap_1.0.12
[16] ggrepel_0.9.1 ggplot2_3.3.3
loaded via a namespace (and not attached):
[1] GOstats_2.56.0 backports_1.2.1 BiocFileCache_1.14.0
[4] plyr_1.8.6 GSEABase_1.52.1 splines_4.0.4
[7] BiocParallel_1.24.1 crosstalk_1.1.1 amap_0.8-18
[10] digest_0.6.27 invgamma_1.1 htmltools_0.5.1.1
[13] GO.db_3.12.1 SQUAREM_2021.1 fansi_0.4.2
[16] magrittr_2.0.1 checkmate_2.0.0 memoise_2.0.0
[19] BSgenome_1.58.0 base64url_1.4 Biostrings_2.58.0
[22] annotate_1.68.0 systemPipeR_1.24.3 bdsmatrix_1.3-4
[25] askpass_1.1 prettyunits_1.1.1 jpeg_0.1-8.1
[28] colorspace_2.0-0 apeglm_1.12.0 blob_1.2.1
[31] rappdirs_0.3.3 xfun_0.21 dplyr_1.0.4
[34] crayon_1.4.1 RCurl_1.98-1.2 jsonlite_1.7.2
[37] graph_1.68.0 genefilter_1.72.1 brew_1.0-6
[40] survival_3.2-7 VariantAnnotation_1.36.0 glue_1.4.2
[43] gtable_0.3.0 zlibbioc_1.36.0 XVector_0.30.0
[46] webshot_0.5.2 DelayedArray_0.16.1 V8_3.4.0
[49] Rgraphviz_2.34.0 scales_1.1.1 mvtnorm_1.1-1
[52] DBI_1.1.1 edgeR_3.32.1 miniUI_0.1.1.1
[55] Rcpp_1.0.6 emdbook_1.3.12 xtable_1.8-4
[58] progress_1.2.2 bit_4.0.4 rsvg_2.1
[61] truncnorm_1.0-8 AnnotationForge_1.32.0 htmlwidgets_1.5.3
[64] httr_1.4.2 gplots_3.1.1 ellipsis_0.3.1
[67] pkgconfig_2.0.3 XML_3.99-0.5 sass_0.3.1
[70] dbplyr_2.1.0 locfit_1.5-9.4 utf8_1.1.4
[73] tidyselect_1.1.0 rlang_0.4.10 manipulateWidget_0.10.1
[76] later_1.1.0.1 AnnotationDbi_1.52.0 munsell_0.5.0
[79] tools_4.0.4 cachem_1.0.4 cli_2.3.0
[82] generics_0.1.0 RSQLite_2.2.3 evaluate_0.14
[85] stringr_1.4.0 fastmap_1.1.0 yaml_2.2.1
[88] knitr_1.31 bit64_4.0.5 caTools_1.18.1
[91] purrr_0.3.4 RBGL_1.66.0 mime_0.10
[94] xml2_1.3.2 biomaRt_2.46.3 compiler_4.0.4
[97] rstudioapi_0.13 curl_4.3 png_0.1-7
[100] tibble_3.0.6 geneplotter_1.68.0 bslib_0.2.4
[103] stringi_1.5.3 GenomicFeatures_1.42.1 lattice_0.20-41
[106] Matrix_1.3-2 vctrs_0.3.6 pillar_1.5.0
[109] lifecycle_1.0.0 jquerylib_0.1.3 irlba_2.3.3
[112] data.table_1.14.0 bitops_1.0-6 httpuv_1.5.5
[115] rtracklayer_1.50.0 R6_2.5.0 latticeExtra_0.6-29
[118] hwriter_1.3.2 promises_1.2.0.1 ShortRead_1.48.0
[121] KernSmooth_2.23-18 MASS_7.3-53.1 gtools_3.8.2
[124] assertthat_0.2.1 openssl_1.4.3 Category_2.56.0
[127] rjson_0.2.20 withr_2.4.1 GenomicAlignments_1.26.0
[130] batchtools_0.9.15 Rsamtools_2.6.0 GenomeInfoDbData_1.2.4
[133] hms_1.0.0 coda_0.19-4 DOT_0.1
[136] rmarkdown_2.7 GreyListChIP_1.22.0 ashr_2.2-47
[139] mixsqp_0.3-43 bbmle_1.0.23.1 numDeriv_2016.8-1.1
[142] shiny_1.6.0
954 != 738+437 ...
Note: crossposted at Bioconductor, but no answer received so far.
Independent filtering?
I fetched the results from the DESeq object with independent filtering set to FALSE.
res <- results(dds, independentFiltering=F, contrast=c("Tissue","MCF7","BT474"))Yes, this is why I am asking. Does DiffBind do the same?
Thanks. I'm not sure what are the defaults of DiffBind, but DESEQ results with independentFiltering=T does not coincide with DiffBind results. Using the same code as above :
Even more importantly, the fold changes and (the p-values) between the two runs are different (so its certainly not an issue of independent filtering)
You can see from the normalized counts that this is the same gene. It has different fold values at the DESeq and at the DiffBind results.
I have opened a new post on that at BioConductor.
Please don't delete posts that have already received comments/answers.