Contents

[!TIP] For minimal setup command, go to Minimal setup section

1. Overview
2. Minimal setup
   1. With BED input
   2. With GTF/GFF3 input
3. Output
   1. ${prefix}.toga.transcripts.bed
   2. ${prefix}.toga.isoforms.tsv
   3. ${prefix}.toga.U12introns.bed
   4. ${prefix}.toga.genes.bed
   5. ${prefix}.sleasy.exons.2bit
   6. rejected_items.tsv
   7. Immunoglobulin annotation files
4. Advanced settings
   1. Isoform filtering
   2. Minimal exon and intron lengths
   3. Nonsense-mediated decay filter

Overview

Given a reference genome sequence and a minimal file with coding transcripts’ coordinates, all the remaining input files can be generate with the TOGA2 prepare-input mode.

Minimal setup

By default, prepare-input requires two named arguments to proceed:

• --ref_2bit is a path to the reference genome sequence, in 2bit format
• --ref_annot is a path to the source reference annotation file, either in BED or in GTF/GFF3 format

The exact minimal settings differ depending on the --ref_annot file format. In addition, running prepare-input with NCBI-style GTF/GFF3 input yields additional input files (see Output section for more details).

With BED input

toga2.py prepare-input --ref_2bit my_ref.2bit --ref_annot my_ref.bed

With GTF/GFF3 input

toga2.py prepare-input --ref_2bit my_ref.2bit --ref_annot my_ref.gtf --annot_format gtf

, or, if a GFF3 file is provided:

toga2.py prepare-input --ref_2bit my_ref.2bit --ref_annot my_ref.gff3 --annot_format gff

Output

By default, the results are saved to `` directory. A custom output path can be set with --output option.
All file names are prepended with a reference prefix set with `--prefix` option. By default, the prefix is kept empty; nonetheless, it is highly recommended to set `--prefix` to keep track of input files across different references. The breakdown below assumes that the `--prefix` is set to a non-empty value.

${prefix}.toga.transcripts.bed

${prefix}.toga.isoforms.tsv

${prefix}.toga.U12introns.bed

${prefix}.toga.genes.bed

${prefix}.sleasy.exons.2bit

rejected_items.tsv

Immunoglobulin annotation files

Advanced settings

Isoform filtering

[!IMPORTANT] External isoform mapping is (currently) not supported if GTF/GFF3 input is provided. Setting both --ref_isoforms and --annot_format gtf/gff causes error-exit with the following error message:

Ref annotation format was set to gtf/gff, which is incompatible with the external isoform mapping. Please remove the isoform file from the settings or provide a BED annotation file instead.

Minimal exon and intron lengths

Nonsense-mediated decay filter

Intron length filter for IntronIC

Intron length filter for CESAR2 profiles