This specification describes allowed input and required output of method_workflows to ensure that the benchmarking_workflows have access to files of the same format from each participant.
Outputs depend on features available for the participant (i.e. not all participants perform de novo identification of PAS) and can be grouped into four categories:
- identification benchmarking event:
- BED file with identified poly(A) sites with single nucleotide resolution
- absolute quantification benchmarking event:
- BED file with identified unique poly(A) sites and TPM values for each site
- relative quantification benchmarking event:
- BED file with identified unique poly(A) sites and relative usage values for each site
- differential usage benchmarking event (not yet implemented)
- TSV file with gene ID and significance of differential PAS usage
Inputs to method workflows are provided by APAeval.
| # | Format | Link | Example data | Description |
|---|---|---|---|---|
| 1 | BAM | Specification | - | BAM file with mapped RNA seq reads Pre-processing done with nfcore-rnaseq |
| 2 | GTF | Specification | Link | GTF file with genome annotation including 3' UTRs |
| OUTCODE | Format | Link | Example data |
|---|---|---|---|
| 01 | BED | Specification | Link |
| 02 | BED | Specification | Link |
| 03 | TSV | Wikipedia | Not yet implemented |
| 04 | BED | Specification | Link |
This BED file contains single-nucleotide position of poly(A) sites identified by the participant.
Fields:
- chrom - the name of the chromosome
- chromStart - the starting position of the feature in the chromosome; this corresponds to the last nucleotide just upstream of the cleavage and polyadenylation reaction; the starting position is 0-based, i.e. the first base on the chromosome is numbered 0
- chromEnd - the ending position of the feature in the chromosome; as identified PAS are single-nucleotide, the ending position is equal to
chromStart + 1 - name - defines the name of the identified poly(A) site
- score - not used, leave as "."
- strand - defines the strand; either "." (=no strand) or "+" or "-".
This BED file contains positions of unique poly(A) sites with TPM values for each identified site in the score column.
Fields:
- chrom - the name of the chromosome
- chromStart - the starting position of the feature in the chromosome; this corresponds to the last nucleotide just upstream of the cleavage and polyadenylation reaction; the starting position is 0-based, i.e. the first base on the chromosome is numbered 0
- chromEnd - the ending position of the feature in the chromosome; as identified PAS are single-nucleotide, the ending position is equal to
chromStart + 1 - name - defines the name of the identified poly(A) site
- score - TPM value for the identified site
- strand - defines the strand; either "." (=no strand) or "+" or "-".
This TSV file contains two columns:
- gene ID
- significance of differential PAS usage
Column names should not be added to the file.
This BED file contains positions of unique poly(A) sites with relative usage values for each identified site in the score column. The relative usage values must be fractional for each poly(A) site (e.g. fraction used vs other sites in the same terminal exon).
Fields:
- chrom - the name of the chromosome
- chromStart - the starting position of the feature in the chromosome; this corresponds to the last nucleotide just upstream of the cleavage and polyadenylation reaction; the starting position is 0-based, i.e. the first base on the chromosome is numbered 0
- chromEnd - the ending position of the feature in the chromosome; as identified PAS are single-nucleotide, the ending position is equal to
chromStart + 1 - name - defines the name of the identified poly(A) site
- score - relative usage value for the identified site (between 0-1)
- strand - defines the strand; either "." (=no strand) or "+" or "-".
For naming conventions please refer to the method workflow README