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  • Can I get read counts by doing this???

    I want to quantify the reads I have from several .bam files from Tophat.

    I used two different annotations(one at a time):
    1: CuffMerge generated GTF file;
    2: UCSC genome annotation
    I made a transcript data base by using Bioconductor package "GenomicFeature" function "makeTranscitpDbFromGFF" or "makeTranscitpDbFromUCSC"

    Then I make a object that store the exon or transcripts information, by using Rsamtools package function "exonsBy" or "transcriptsBy"

    Then I do the overlap between my .bam file and the previous object ("countOverlaps")

    Anyone see any problems by doing what I did?

    I took this as a reference:

    Please let me know what could go wrong with this~

    Thanks a lot!

  • #2
    Be careful using countOverlaps:

    Code:
    library(IRanges)
    subject <- IRanges(c(1, 4, 9), c(5, 7, 10))
    query <- IRanges(c(2, 2, 10), c(2, 3, 12))
    #If this were RNAseq data, the output should be 0 0 1
    countOverlaps(query, subject)
    You'll see that the counts would be wrong for RNAseq data. You might just use featureCounts or htseq-counts.

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