Miso Index creator not working with ensEMBL GFF files

[wariobrega]

Hello everybody.

I am having troubles using the MISO pipeline for finding alternative first exons on my reads dataset.

for making MISO work, an index must be created by the index_gff.py script which comes with the pipelne. According to the MISO webiste, the input should be like this one:

chr1 SE gene 4772649 4775821 . - . ID=chr1:4775654:4775821:-@chr1:4774032:4774186:-@chr1:4772649:4772814:-;Name=chr1:4775654:4775821:-@chr1:4774032:4774186:-@chr1:4772649:4772814:-
chr1 SE mRNA 4772649 4775821 . - . ID=chr1:4775654:4775821:-@chr1:4774032:4774186:-@chr1:4772649:4772814:-.A;Parent=chr1:4775654:4775821:-@chr1:4774032:4774186:-@chr1:4772649:4772814:-
chr1 SE mRNA 4772649 4775821 . - . ID=chr1:4775654:4775821:-@chr1:4774032:4774186:-@chr1:4772649:4772814:-.B;Parent=chr1:4775654:4775821:-@chr1:4774032:4774186:-@chr1:4772649:4772814:-
chr1 SE exon 4775654 4775821 . - . ID=chr1:4775654:4775821:-@chr1:4774032:4774186:-@chr1:4772649:4772814:-.A.up;Parent=chr1:4775654:4775821:-@chr1:4774032:4774186:-@chr1:4772649:4772814:-.A
chr1 SE exon 4774032 4774186 . - . ID=chr1:4775654:4775821:-@chr1:4774032:4774186:-@chr1:4772649:4772814:-.A.se;Parent=chr1:4775654:4775821:-@chr1:4774032:4774186:-@chr1:4772649:4772814:-.A
chr1 SE exon 4772649 4772814 . - . ID=chr1:4775654:4775821:-@chr1:4774032:4774186:-@chr1:4772649:4772814:-.A.dn;Parent=chr1:4775654:4775821:-@chr1:4774032:4774186:-@chr1:4772649:4772814:-.A
chr1 SE exon 4775654 4775821 . - . ID=chr1:4775654:4775821:-@chr1:4774032:4774186:-@chr1:4772649:4772814:-.B.up;Parent=chr1:4775654:4775821:-@chr1:4774032:4774186:-@chr1:4772649:4772814:-.B
chr1 SE exon 4772649 4772814 . - . ID=chr1:4775654:4775821:-@chr1:4774032:4774186:-@chr1:4772649:4772814:-.B.dn;Parent=chr1:4775654:4775821:-@chr1:4774032:4774186:-@chr1:4772649:4772814:-.B

Instead, a normal GFF file downloaded from ensEMBL looks like this

11 protein_coding exon 31158398 31158571 . + . gene_id "ENSDARG00000091602"; transcript_id "ENSDART00000122756"; exon_number "2"; gene_name "CR790368.1"; gene_biotype "protein_coding"; transcript_name "CR790368.1-201";
11 protein_coding CDS 31158398 31158571 . + 2 gene_id "ENSDARG00000091602"; transcript_id "ENSDART00000122756"; exon_number "2"; gene_name "CR790368.1"; gene_biotype "protein_coding"; transcript_name "CR790368.1-201"; protein_id "ENSDARP00000111013";
11 protein_coding exon 31161221 31161385 . + . gene_id "ENSDARG00000091602"; transcript_id "ENSDART00000122756"; exon_number "3"; gene_name "CR790368.1"; gene_biotype "protein_coding"; transcript_name "CR790368.1-201";
11 protein_coding CDS 31161221 31161385 . + 2 gene_id "ENSDARG00000091602"; transcript_id "ENSDART00000122756"; exon_number "3"; gene_name "CR790368.1"; gene_biotype "protein_coding"; transcript_name "CR790368.1-201"; protein_id "ENSDARP00000111013";
11 protein_coding exon 31161542 31162705 . + . gene_id "ENSDARG00000091602"; transcript_id "ENSDART00000122756"; exon_number "4"; gene_name "CR790368.1"; gene_biotype "protein_coding"; transcript_name "CR790368.1-201";
11 protein_coding CDS 31161542 31162705 . + 2 gene_id "ENSDARG00000091602"; transcript_id "ENSDART00000122756"; exon_number "4"; gene_name "CR790368.1"; gene_biotype "protein_coding"; transcript_name "CR790368.1-201"; protein_id "ENSDARP00000111013";
11 protein_coding exon 31165694 31165819 . + . gene_id "ENSDARG00000091602"; transcript_id "ENSDART00000122756"; exon_number "5"; gene_name "CR790368.1"; gene_biotype "protein_coding"; transcript_name "CR790368.1-201";
11 protein_coding CDS 31165694 31165819 . + 2 gene_id "ENSDARG00000091602"; transcript_id "ENSDART00000122756"; exon_number "5"; gene_name "CR790368.1"; gene_biotype "protein_coding"; transcript_name "CR790368.1-201"; protein_id "ENSDARP00000111013";
11 protein_coding exon 31167191 31167313 . + . gene_id "ENSDARG00000091602"; transcript_id "ENSDART00000122756"; exon_number "6"; gene_name "CR790368.1"; gene_biotype "protein_coding"; transcript_name "CR790368.1-201";
11 protein_coding CDS 31167191 31167313 . + 2 gene_id "ENSDARG00000091602"; transcript_id "ENSDART00000122756"; exon_number "6"; gene_name "CR790368.1"; gene_biotype "protein_coding"; transcript_name "CR790368.1-201"; protein_id "ENSDARP00000111013";
11 protein_coding exon 31167542 31167667 . + . gene_id "ENSDARG00000091602"; transcript_id "ENSDART00000122756"; exon_number "7"; gene_name "CR790368.1"; gene_biotype "protein_coding"; transcript_name "CR790368.1-201";
11 protein_coding CDS 31167542 31167667 . + 2 gene_id "ENSDARG00000091602"; transcript_id "ENSDART00000122756"; exon_number "7"; gene_name "CR790368.1"; gene_biotype "protein_coding"; transcript_name "CR790368.1-201"; protein_id "ENSDARP00000111013";
11 protein_coding exon 31168516 31168641 . + . gene_id "ENSDARG00000091602"; transcript_id "ENSDART00000122756"; exon_number "8"; gene_name "CR790368.1"; gene_biotype "protein_coding"; transcript_name "CR790368.1-201";
11 protein_coding CDS 31168516 31168641 . + 2 gene_id "ENSDARG00000091602"; transcript_id "ENSDART00000122756"; exon_number "8"; gene_name "CR790368.1"; gene_biotype "protein_coding"; transcript_name "CR790368.1-201"; protein_id "ENSDARP00000111013";
11 protein_coding exon 31168791 31168945 . + . gene_id "ENSDARG00000091602"; transcript_id "ENSDART00000122756"; exon_number "9"; gene_name "CR790368.1"; gene_biotype "protein_coding"; transcript_name "CR790368.1-201";
11 protein_coding CDS 31168791 31168942 . + 2 gene_id "ENSDARG00000091602"; transcript_id "ENSDART00000122756"; exon_number "9"; gene_name "CR790368.1"; gene_biotype "protein_coding"; transcript_name "CR790368.1-201"; protein_id "ENSDARP00000111013";
11 protein_coding stop_codon 31168943 31168945 . + 0 gene_id "ENSDARG00000091602"; transcript_id "ENSDART00000122756"; exon_number "9"; gene_name "CR790368.1"; gene_biotype "protein_coding"; transcript_name "CR790368.1-201";

so the indexer is not recognizing some part, as described by the error log:
[CODEWARNING: Invalid attributes string: gene_id "ENSDARG00000074181"; transcript_id "ENSDART00000057160"; exon_number "1"; gene_name "LRRC24 (2 of 2)"; gene_biotype "protein_coding"; transcript_name "CT027690.1-201";[/CODE]

I would like to change the script (which is in python) but since I don't know the indexing criterias I don't know how to do it. Has some already done this, or know a way to make this script indexing work?

thank you very much,

daniele

[dadada4ever]

The normal GFF file you posted below provided by ENSEMBL is gtf format, you should use gtf2gff3.pl to convert gtf to gff file first, then index the gff file which is successfully converted.
It's mentioned in MISO manual:

http://genes.mit.edu/burgelab/miso/scripts/gtf2gff3.pl