To Whom It May Concern,

I am currently working on a project examining differential expression in RNA-seq data with DEXseq and I have encountered a few flags when using the dexseq_prepare_annotation.py script to interpret my .gtf file. My annotation file was generated by NCBI and I initially had to change 'Parent' to 'gene_id' in the attribute column (please see below). However now I am getting another flag regarding 'transcript_id' but I don't understand why? Any insight as to how I can modify the format to meet the criteria for proper .gtf input would be greatly appreciated.

Thank you,

A. Romney

error:

(env2)arom2:~:1015 > python /vol/apps/user/stow/R-3.2.1/lib64/R/library/DEXSeq/python_scripts/dexseq_prepare_annotation.py fhet_prep.gtf Fhet_final.gff
Traceback (most recent call last):
  File "/vol/apps/user/stow/R-3.2.1/lib64/R/library/DEXSeq/python_scripts/dexseq_prepare_annotation.py", line 55, in <module>
    exons[f.iv] += ( f.attr['gene_id'], f.attr['transcript_id'] )
KeyError: 'transcript_id'

Here is a sub-section of the gtf file I am using to show you how the gene_id and transcript_id are identified.

NW_012224401.1    Gnomon    exon    71785    71884    .    +    .    ID=id44;gene_id=rna2;Dbxref=GeneID:105915271,Genbank:XM_012849362.1;gbkey=mRNA;gene=csf1r;product=colony stimulating factor 1 receptor%2C transcript variant X1;transcript_id=XM_012849362.1
NW_012224401.1    Gnomon    exon    72804    72915    .    +    .    ID=id45;gene_id=rna2;Dbxref=GeneID:105915271,Genbank:XM_012849362.1;gbkey=mRNA;gene=csf1r;product=colony stimulating factor 1 receptor%2C transcript variant X1;transcript_id=XM_012849362.1
NW_012224401.1    Gnomon    exon    76564    76791    .    +    .    ID=id46;gene_id=rna2;Dbxref=GeneID:105915271,Genbank:XM_012849362.1;gbkey=mRNA;gene=csf1r;product=colony stimulating factor 1 receptor%2C transcript variant X1;transcript_id=XM_012849362.1
NW_012224401.1    Gnomon    mRNA    62183    76791    .    +    .    ID=rna3;gene_id=gene2;Dbxref=GeneID:105915271,Genbank:XM_012849437.1;Name=XM_012849437.1;gbkey=mRNA;gene=csf1r;product=colony stimulating factor 1 receptor%2C transcript variant X2;transcript_id=XM_012849437.1
NW_012224401.1    Gnomon    exon    62183    62306    .    +    .    ID=id47;gene_id=rna3;Dbxref=GeneID:105915271,Genbank:XM_012849437.1;gbkey=mRNA;gene=csf1r;product=colony stimulating factor 1 receptor%2C transcript variant X2;transcript_id=XM_012849437.1
NW_012224401.1    Gnomon    exon    62565    62634    .    +    .    ID=id48;gene_id=rna3;Dbxref=GeneID:105915271,Genbank:XM_012849437.1;gbkey=mRNA;gene=csf1r;product=colony stimulating factor 1 receptor%2C transcript variant X2;transcript_id=XM_012849437.1
NW_012224401.1    Gnomon    exon    63886    64173    .    +    .    ID=id49;gene_id=rna3;Dbxref=GeneID:105915271,Genbank:XM_012849437.1;gbkey=mRNA;gene=csf1r;product=colony stimulating factor 1 receptor%2C transcript variant X2;transcript_id=XM_012849437.1
NW_012224401.1    Gnomon    exon    64260    64547    .    +    .    ID=id50;gene_id=rna3;Dbxref=GeneID:105915271,Genbank:XM_012849437.1;gbkey=mRNA;gene=csf1r;product=colony stimulating factor 1 receptor%2C transcript variant X2;transcript_id=XM_012849437.1
NW_012224401.1    Gnomon    exon    64630    64766    .    +    .    ID=id51;gene_id=rna3;Dbxref=GeneID:105915271,Genbank:XM_012849437.1;gbkey=mRNA;gene=csf1r;product=colony stimulating factor 1 receptor%2C transcript variant X2;transcript_id=XM_012849437.1
NW_012224401.1    Gnomon    exon    64845    65010    .    +    .    ID=id52;gene_id=rna3;Dbxref=GeneID:105915271,Genbank:XM_012849437.1;gbkey=mRNA;gene=csf1r;product=colony stimulating factor 1 receptor%2C transcript variant X2;transcript_id=XM_012849437.1

Thanks Alejandro,

I was able to revise the NCBI annotation file I am using and can now also get it to work with the prep_annotation script.

Thanks NCBI...