ANNOVAR to annotate variants called with the SAMtools pipeline

[MGCBrown]

Hello,

I'm new to this forum and bioinformatics for that matter.

I'm studying functional genomics and ecology of hybridization in flying squirrels, with the goal of identifying SNPs between two flying squirrel species, and profiling hybrid individuals based on a handful of those variable markers.

I've de novo assembled a flying squirrel transcriptome with Trinity, and have called SNPs using the SAMtools variant calling pipeline (mpileup etc).

I now would like to use ANNOVAR to annotate the SNPs that SAMtools has called. ANNOVAR provides detailed documentation on annotating SNPs based of human genomic databases. I believe that the closest open-source annotated genome to my study species in the thirteen-lined ground squirrel (SpeTri2), followed by the mouse (mm9).

I'm confused as to what databases from the SpeTri2 annotated assembly I should be downloading and using in ANNOVAR, as I noticed that in the ANNOVAR quick-start guide, the directions called for many human genomic databases.

Any info would be great.

Cheers.

[MGCBrown]

It seems that a more pressing issue is that the SAMtools variant calling pipeline has created vCalendar files and not VCF (variant call format) files.

I followed this pipeline:
samtools mpileup -uf ref.fa aln1.bam aln2.bam | bcftools view -bvcg - > var.raw.bcf
bcftools view var.raw.bcf | vcfutils.pl varFilter -D100 > var.flt.vcf

Does anyone know why I would end up with a vCalendar file? I'm guessing it is a completely different format from a VCF file?

[GenoMax]

On your computer .vcf extension may be associated with the calendar app.

If you use a programmer's editor (Notepad++ for Windows or BBEdit/Textedit for macOS) you should be able to view the vcf file (File --> Open --> FInd the file). If all looks ok then you would want to remove that extension association.

[MGCBrown]

Thanks for the tips!

I've opened the .vcf file in Notepad++. I've copied two components of my .vcf file below as one of the components looks OK but the other, not so much. Do you think someone went wrong during the SAMtools pipeline?


##contig=<ID=TRINITY_DN393704_c0_g1_i1,length=379,IDX=665834>
##contig=<ID=TRINITY_DN393738_c0_g1_i1,length=262,IDX=665835>
##contig=<ID=TRINITY_DN393713_c0_g1_i1,length=215,IDX=665836>
##contig=<ID=TRINITY_DN393739_c0_g1_i1,length=208,IDX=665837>
##contig=<ID=TRINITY_DN393753_c0_g1_i1,length=358,IDX=665838>
##contig=<ID=TRINITY_DN393757_c0_g1_i1,length=304,IDX=665839>
##contig=<ID=TRINITY_DN393784_c0_g1_i1,length=556,IDX=665840>
##contig=<ID=TRINITY_DN393725_c0_g1_i1,length=221,IDX=665841>
##contig=<ID=TRINITY_DN393774_c0_g1_i1,length=245,IDX=665842>
##contig=<ID=TRINITY_DN393710_c0_g1_i1,length=382,IDX=665843>
##contig=<ID=TRINITY_DN393772_c0_g1_i1,length=276,IDX=665844>
##contig=<ID=TRINITY_DN393785_c0_g1_i1,length=531,IDX=665845>
##contig=<ID=TRINITY_DN393793_c0_g1_i1,length=274,IDX=665846>
##contig=<ID=TRINITY_DN393728_c0_g1_i1,length=291,IDX=665847>
##contig=<ID=TRINITY_DN393767_c0_g1_i1,length=226,IDX=665848>
##contig=<ID=TRINITY_DN393716_c0_g1_i1,length=222,IDX=665849>



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