Can't get bedtools intersect with split option to work with BED12 file with multiple blocks

[Saj Batra]

Hi All,
I expect the following intersection to produce no overlaps as the probe sequence does not overlap with any of the blocks. However I obtain 25 bases overlap. What am I doing wrong?
I'm using:
Tool: bedtools intersect (aka intersectBed)
Version: v2.26.0

Here are my input BED12 files:
cat toy_probe.bed
chr1 1000075 1000100 probe 1084 +1 1000075 1000075 255,0,0 1 25 0

cat toy_mrna.bed
chr1 1000000 1002000 mrna 1084 +1 1000000 1000000 255,0,0 5 40,400,160,200,300, 0,300,1000,1200,1700

/nfs/netaffx/tools/bedtools/bedtools2/bin/bedtools intersect -split -wao -a toy_probe.bed -b toy_mrna.bed
chr1 1000075 1000100 probe 1084 +1 1000075 1000075 255,0,0 1 25 0 chr1 1000000 1002000 mrna 1084 +1 1000000 1000000 255,0,0 5 40,400,160,200,300, 0,300,1000,1200,1700 25


Since probe only has 1 block, I've also tried running it with probe as a BED6 file as follows:
cat toy_probe2.bed
chr1 1000075 1000100 probe 1084 +1

/nfs/netaffx/tools/bedtools/bedtools2/bin/bedtools intersect -split -wao -a toy_probe2.bed -b toy_mrna.bed
chr1 1000075 1000100 probe 1084 +1 chr1 1000000 1002000 mrna 1084 +1 1000000 1000000 255,0,0 5 40,400,160,200,300, 0,300,1000,1200,1700 25

I'm also attaching a picture of the mrna, probe both aligned in the UCSC Genome Browser (HG38). The picture shows that there is no overlap between the mrna and probe.
Any ideas as to how to get this to not produce any overlap?

Thanks, Sajeev

[Saj Batra]

FYI -  I can now get the split option to work if I change the strand to "+"  (or "-" when necessary).  

However intersect seems to be ignoring strand information and I get overlaps for different strands.  

Anybody now how to fix that?  It seems like I need another filtering step?

Thanks

[Aaron Quinlan]

Dear Sajeev,

If I understand correctly, the -s option will solve your issue.

Best,

Aaron

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[Saj Batra]

Thanks Aaron, -s has solved my issue.

Cheers, Sajeev

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