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Old 11-01-2018, 01:12 PM   #1
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Default featureCounts - order of reads in output file


We use featureCounts a lot and firstly thanks to Dr. Shi and R. Liao for a great tool.

I had a question about featureCounts output, specifically the order of the reads in the output file. So, I ran featureCounts on a bam file against a given GTF, with two summarization options (-t exon and -t gene) trying to compare the results side-by-side, using following options:
/featureCounts --primary -R CORE -F GTF -s 0 -Q 0 -t exon -g gene_id -T 8 -a Homo_sapiens.GRCh38.81.gtf -p -o test1.exon.featureCounts test1.Aligned.out.bam
/featureCounts --primary -R CORE -F GTF -s 0 -Q 0 -t gene -g gene_id -T 8 -a Homo_sapiens.GRCh38.81.gtf -p -o test1.gene.featureCounts test1.Aligned.out.bam

I found that the order of reads in the output files (the .bam.featureCounts file that is at the read level) were different. The total number of lines were the same.

Q1: Is there is an option to keep the order in the output the same?
Q2: I also observed that some reads have multiple lines of output for multi-mapping (from multiple multi-mapping hits in the bam?) and sometimes at different places in the file (so, collapsing contiguous lines using uniq does not alleviate it). Is it possible to tackle this somehow? Eg:
Read1 Unassigned_MultiMapping -1 NA
Read1 Unassigned_MultiMapping -1 NA
Read1 Unassigned_MultiMapping -1 NA

The bam file was the output of STAR aligner on paired end RNASeq data (Unsorted).

Many thanks for your replies in advance.
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