I'm working on Coli MG1655, after i have aligned the reads to reference, i checked in IGV if alignments are coherent to gene expression but i noticed that despite some genes are mapped on forward strand, the reads has been aligned to the reverse strand.
i used samtools to divide reads mapped on forward and reverse strand in this way:
samtools view -F 16 -b -o forward_strand.bam in.bam
samtools view -f 16 -b -o reverse_strand.bam in.bam
I am attaching an example of groL gene, mapped to the forward strand, but reads actually mapped on reverse strand.
Why? what am i missing?
In the image, from top to bottom: gff annotation, coverage reverse, coverage forward, mapped reads to reverse strand.