using −−donotsort command with Featurecounts

[R adele]

Hello I did  paired-ended RNA seq .  I now want to do the read counts using Featurecounts. I had the following questions 

1) Can someone explain the −−donotsort option on Featurecounts.  The Featurecounts manual defines it as :  −−donotsort : "If specified, paired end reads will not be re-ordered even if reads from the same pair were found not to be next to each other in the input"                    I still don't understand what it does exactly ?

2) I wanted to add the −−donotsort option because I'm afraid the featurecounts program  when  it does the read counts it would not be able to properly account for paired reads that are not close in proximity due to being a part of a duplication or deletion or  being chimeric reads. Is it ok to  add the  −−donotsort option for this reason? Below is the command line I used: 

/hpf/tools/centos6/subread/1.5.3/bin/featureCounts -a /hpf/projects/deon/sequencing_repository/raw_fastq/RNA_bobby/bam/gencode.v19.annotation.gtf -o family_13_01_RNA-seq.counts -g gene_name -p -s 2 -C --donotsort  D4775_R312_BAligned.sortedByCoord.out.bam D4754_R311_BAligned.sortedByCoord.out.bam D4776_R310Aligned.sortedByCoord.out.bam D4777_R307Aligned.sortedByCoord.out.bam D4778_R308BAligned.sortedByCoord.out.bam D4828_R309_BAligned.sortedByCoord.out.bam 

[Yang LIAO]

The "--donotsort" option for featureCounts is for dealing with the SAM or BAM files where two reads in the same pair don't have the same read name. Some read aligners have this problem by not removing "/1" or "/2" from the end of the read names, making featureCounts treat the two reads in the same pair as unrelated reads. By specifying this option, featureCounts doesn't try to pair up the reads, but treat every next two reads as in the same pair.

The commonly used aligners all don't have this issue. Unless you have found this issue from the SAM or BAM file, you shouldn't use this option.

[R adele]

Thanks Yang Liao  for answering my question