Running two bedtools commands in series produces error "has non positional records"

Question

[Lina F]

Hello all,

I have three samples, generated Illumina PE reads for each of them, and then mapped the reads against the reference to produce three alignments. I called SNPs and now I would like to determine the subset of SNPs that are present in sample A and B but not in C. I ran bedtools as follows:

bedtools intersect -a A.vcf -b B.vcf > A_and_B.vcf
bedtools subtract -a A_and_B.vcf -b C.vcf

Unfortunately, I get the following error message:

ERROR: file A_and_B.vcf has non positional records, which are only valid for the groupBy tool.

I checked that all my files are tab-delimited and I also ran sort -k1 on the A_and_B.vcf file but that didn't affect the output.

My questions are:

1. How do I string these two bedtools commands together?
2. Is there a better way to do this?

Thanks for any suggestions!

[Nitin Narwade]

you can try something like this.

 bedtools intersect -a A.vcf -b B.vcf | bedtools subtract -a stdin -b C.vcf

[Lina F]

Thank you for the suggestion! Unfortunately, this didn't work for me either :-(

Answer 1

[finswimmer]

There is an -header option for bedtools intersect which include the header in the output file.

-header Print the header from the A file prior to results.

fin swimmer

[Lina F]

Thanks for the tip, this worked and let's me avoid copying things manually!

[ATpoint]

You should move that to answer, because that is exactly the point.

Answer 2

[Lina F]

I found a potential workaround. First, I upgraded bedtools from v2.26.0 to v2.27.1. This changed the error message that was reported and made it easier to interpret:

Error: unable to open file or unable to determine types for file A_and_B.vcf

- Please ensure that your file is TAB delimited (e.g., cat -t FILE).
- Also ensure that your file has integer chromosome coordinates in the
  expected columns (e.g., cols 2 and 3 for BED).

I took another look at the output file and I noticed while it is tab-delimited, it is missing the header (which specifies that the input is in VCF format). I manually copied the header of one of the input files to the intermediate output file. Now my second command, bedtools subtract, works.

This seems like a way forward, with the caveat being that the header I manually copied to the intermediate file is not entirely correct.

If there is a better way to do this I'd love to hear about it!