interpretation of gstacks output file using Stacks v2 Beta version

[Carol]

Hi everyone!!

Following the recent release of STACKS v2 (with its interesting new features), I gave it a try by running gstacks for one of my paired-end aligned samples. 

My aim was to test different .bam files resulting from different aligners: bowtie2 (--very-sensitive-local), bwa mem (default), and gsnap (default, 2, 3, and 4 max mismatches allowed)

The code I used was: 

gstacks -B ./bowtie.rg_sorted.bam -O ./bowtie --pair -t 30 --details -m 3 --max-insert 600

gstacks -B ./bwa.rg_sorted.bam -O ./bwa --pair -t 30 --details -m 3 --max-insert 600

gstacks -B ./gsnap8mm.rg_sorted.bam -O ./gsnap8 --pair -t 30 --details -m 3 --max-insert 600

gstacks -B ./gsnap4mm.rg_sorted.bam -O ./gsnap4 --pair -t 30 --details -m 3 --max-insert 600

gstacks -B ./gsnap3mm.rg_sorted.bam -O ./gsnap3 --pair -t 30 --details -m 3 --max-insert 600

gstacks -B ./gsnap2mm.rg_sorted.bam -O ./gsnap2 --pair -t 30 --details -m 3 --max-insert 600

The outputfiles are:

gstacks.fa.gz

gstacks.details.gz

gstacks.vcf.gz

gstacks.log

(for each of my bam files)

I was expecting to be able to see how homozygotes and heterozygotes were called for each of my test alignments, but since in this new version there is not XXX.snps.tvs file I was unable to. I though similar information could be found in the vcf file, but column #ALT is completely empty, same for #ID, #QUAL and #FILTER.

Where can I now find the analogous output information of what previously was pstacks??

How can I see which loci are homozygous or heterozygous in my sample??

Where is the RADtags catalog? all the samples used for an analysis will be included in the catalog??...

I understand that the version was recently released, but a bit more of information about how to interpret output files will be of great help!!

Cheers,

Carol

[Nicolas Rochette]

Hi Carol,

For now it's a beta version so while the core of the pipeline should run well, some parts of the interface are still missing. We're going to release a new version in a couple days that will have the most important ones, and gradually add manual pages etc.

The default way to export genotypes will be to use population's --vcf export option, which I am working on right now. The gstacks.vcf.gz should actually contain the information you want, but it's an internal file that contains lots of information that as a user you probably don't care about and that makes the file harder to work with.

But I don't think all the ALT fields are actually empty; running `zcat gstacks.vcf.gz | awk '$5 != "."'` should show where SNPs have been found. If that doesn't do anything, something else probably went wrong and please let me know.

In any case, thank you for the feedback, we're probably change the file name so as to avoid confusion.
Best,

Nicolas

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Stacks website: http://catchenlab.life.illinois.edu/stacks/
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[Carol Buitrago]

Hi Nicolas,

Thanks for your quick response.

As I stated in my previous message the #ALT column is filled with "."

I tried your command as well as

zcat gstacks.vcf.gz | cut -f5 | sort | uniq -c

65867342 .

  

nothing other than dots "." show up. 

I ran gstacks on a single bam file corresponding to a single alignment, just like if I were trying to run pstacks for a single sample (not sure if to call SNPs bam has to contain more than one sample)

Looking forward for the new version and any other advice you could provide.

Cheers,

Carol

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