cn.mops Error in referencecn.mops Genomic Ranges must be unique

Question

[wes3985]

Hi All, I have come across an error in cn.mops that is not clear how to solve. I haven't used the software before so it may be a quick fix for anyone who has. The function ' referencecn.mops' is returning the following error:

    Error in referencecn.mops(X[, 1], X[, 2], norm = 0, I = c(0.025, 0.5,  :
    Genomic Ranges must be unique. Check "all(isUnique(input))" and remove identical segments.
    Execution halted

As I understand, a 'genomic range object' is a read count matrix that the algorithm converts a BAM file into [1]. So it seems to be suggesting that the BAM files are identical? I have done a head(X[,1]) and head(X[,2]) just to show the inputs if anyone can spot a problem here:

> head(X[,1])
GRanges object with 6 ranges and 1 metadata column:
      seqnames         ranges strand | g14_picard_dedup.sorted.bam
         <Rle>      <IRanges>  <Rle> |                   <numeric>
  [1]     chr1 [12098, 12258]      * |            392.216485975478
  [2]     chr1 [12553, 12721]      * |            542.181612966102
  [3]     chr1 [13331, 13701]      * |            1445.20239302964
  [4]     chr1 [30334, 30503]      * |            267.168641623296
  [5]     chr1 [35045, 35544]      * |            2001.22694079488
  [6]     chr1 [35618, 35778]      * |            488.655598409448

> head(X[,2])
GRanges object with 6 ranges and 1 metadata column:
      seqnames         ranges strand | fc14_picard_dedup.sorted.bam
         <Rle>      <IRanges>  <Rle> |                    <numeric>
  [1]     chr1 [12098, 12258]      * |             408.547928776133
  [2]     chr1 [12553, 12721]      * |             617.262631520462
  [3]     chr1 [13331, 13701]      * |             1989.45078360552
  [4]     chr1 [30334, 30503]      * |             386.747940481754
  [5]     chr1 [35045, 35544]      * |              2282.1358116319
  [6]     chr1 [35618, 35778]      * |             756.136631025392

The row identifiers on the left seem to indicate a genomic range which seems to be identical in both files, though this is what I would expect as the algorithm is supposed to compare local regions for variations in depth to look for cnvs. The numeric columns are a bit variable but again I would expect this for regions of similar coverage from different samples. Am I missing something?

Here is the full code up to where the error occurs:

library(cn.mops)
BAMFiles <- c("somatic.bam", "tumour.bam")
BaiFiles <- c("somatic.bai", "tumour.bai")
segments <- read.table("truseq_hg38.bed",sep="\t",as.is=TRUE)
gr <- GRanges(segments[,1],IRanges(segments[,2],segments[,3]))
X1 <- getSegmentReadCountsFromBAM(BAMFiles,GR=gr,mode="paired", BAIFiles=BaiFiles, parallel=12)
X <- normalizeGenome(X1, normType="poisson")
ref_analysis_14 <- referencecn.mops(X[,1], X[,2],
norm=0,
I = c(0.025, 0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4, 8, 16, 32, 64),
classes = paste0("CN", c(0:8, 16, 32, 64, 128)),
segAlgorithm="DNAcopy")

Answer 1

[wes3985]

For anyone else who comes across this error: I think it is caused by the segments input file having some duplicate lines in it which causes errors downstream in the genomic range objects.

It can be solved as follows:

library(cn.mops)
BAMFiles <- c("somatic.bam", "tumour.bam")
BaiFiles <- c("somatic.bai", "tumour.bai")
segments <- read.table("truseq_hg38.bed",sep="\t",as.is=TRUE)
segments <- unique(segments)
gr <- GRanges(segments[,1],IRanges(segments[,2],segments[,3]))
X1 <- getSegmentReadCountsFromBAM(BAMFiles,GR=gr,mode="paired", BAIFiles=BaiFiles, parallel=12)
X <- normalizeGenome(X1, normType="poisson")
ref_analysis_14 <- referencecn.mops(X[,1], X[,2],
norm=0,
I = c(0.025, 0.5, 1, 1.5, 2, 2.5, 3, 3.5, 4, 8, 16, 32, 64),
classes = paste0("CN", c(0:8, 16, 32, 64, 128)),
segAlgorithm="DNAcopy")