CSAW windowCounts yielding an empty RangedSummarizedExperiment

Question

[blin100]

Hi all, I'm trying to use the csaw package to analyze some ATACseq data, and I'm running into problems with windowCounts()

param<-readParam(minq=50)
bam.files<-c("D:/ATAC_CSAW/BL3_sortedByCoord.bam","D:/ATAC_CSAW/BL2_sortedByCoord.bam")
data<-windowCounts(bam.files, ext=110, width=10, param=param)

Whenever I try and read in my bam files, it runs for a while, and outputs an object, formal class RangedSummarizedExperiment

However, within it, there appears to be no data:

> data
class: RangedSummarizedExperiment 
dim: 0 2 
metadata(6): spacing width ... param final.ext
assays(1): counts
rownames: NULL
rowData names(0):
colnames: NULL
colData names(4): bam.files totals ext rlen

Any ideas as to why this is happening? My bam files are sorted by coordinate, with appropriate .bam.bai files, and not corrupt (samtools quick check passes fine)

sessionInfo()
R version 4.0.1 (2020-06-06)
Platform: x86_64-w64-mingw32/x64 (64-bit)
Running under: Windows >= 8 x64 (build 9200)

Matrix products: default

locale:
[1] LC_COLLATE=English_United States.1252  LC_CTYPE=English_United States.1252    LC_MONETARY=English_United States.1252
[4] LC_NUMERIC=C                           LC_TIME=English_United States.1252    

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

other attached packages:
 [1] edgeR_3.30.3                limma_3.44.3                csaw_1.22.1                 SummarizedExperiment_1.18.1
 [5] DelayedArray_0.14.0         matrixStats_0.56.0          Biobase_2.48.0              GenomicRanges_1.40.0       
 [9] GenomeInfoDb_1.24.0         IRanges_2.22.2              S4Vectors_0.26.1            BiocGenerics_0.34.0        

loaded via a namespace (and not attached):
 [1] locfit_1.5-9.4           Rcpp_1.0.4.6             lattice_0.20-41          prettyunits_1.1.1        Rsamtools_2.4.0         
 [6] Biostrings_2.56.0        assertthat_0.2.1         digest_0.6.25            BiocFileCache_1.12.0     R6_2.4.1                
[11] RSQLite_2.2.0            httr_1.4.1               pillar_1.4.4             zlibbioc_1.34.0          rlang_0.4.6             
[16] GenomicFeatures_1.40.0   progress_1.2.2           curl_4.3                 blob_1.2.1               Matrix_1.2-18           
[21] BiocParallel_1.22.0      stringr_1.4.0            RCurl_1.98-1.2           bit_1.1-15.2             biomaRt_2.44.0          
[26] compiler_4.0.1           rtracklayer_1.48.0       pkgconfig_2.0.3          askpass_1.1              openssl_1.4.1           
[31] tidyselect_1.1.0         tibble_3.0.1             GenomeInfoDbData_1.2.3   XML_3.99-0.3             crayon_1.3.4            
[36] dplyr_1.0.0              dbplyr_1.4.4             GenomicAlignments_1.24.0 bitops_1.0-6             rappdirs_0.3.1          
[41] grid_4.0.1               lifecycle_0.2.0          DBI_1.1.0                magrittr_1.5             stringi_1.4.6           
[46] XVector_0.28.0           ellipsis_0.3.1           generics_0.0.2           vctrs_0.3.1              tools_4.0.1             
[51] bit64_0.9-7              glue_1.4.1               purrr_0.3.4              hms_0.5.3                AnnotationDbi_1.50.0    
[56] BiocManager_1.30.10      memoise_1.1.0

Any help would be greatly appreciated!

Answer 1

[ATpoint]

Did you align with bowtie2, because I am pretty sure the maximum MAPQ bowtie2 assignes is 42, ou set 50 as minimum.

[blin100]

oops, didn't get an email for some reason. Yes, I did align with bowtie. Dear lord, that was a dumb mistake. thanks!

[ATpoint]

Don't worry, these things happen :)

If this solved it please accept the answer to provide closure to this question.