DESCRIPTION
As of this version, variants.gff is our primary variant call file format. The variants.gff file is based on the GFFv3 standard (http://www.sequenceontology.org/gff3.shtml). The GFFv3 standard describes a tab-delimited plain-text file meta-format for describing genomic "features." Each gff file consists of some initial "header" lines supplying metadata, and then a number of "feature" lines providing information about each identified variant.
The GFF Coordinate System
All coordinates in GFF files are 1-based, and all intervals start, end are understood as including both endpoints.
Headers
The variants.gff file begins with a block of metadata headers, which looks like the following:
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##gff-version 3 ##pacbio-variant-version 2.1 ##date Tue Feb 28 17:44:18 2012 ##feature-ontology http://song.cvs.sourceforge.net/*checkout*/song/ontology/sofa.obo?revision=1.12 ##source GenomicConsensus v0.1.0 ##source-commandline callVariants.py --algorithm=plurality aligned_reads.cmp.h5 -r spinach.fasta -o variants.gff ##source-alignment-file /home/popeye/data/aligned_reads.cmp.h5 ##source-reference-file /home/popeye/data/spinach.fasta ##sequence-region EGFR_Exon_23 1 189 ##sequence-header EGFR_Exon_24 1 200
The source and source-commandline describe the name and version of the software generating the file. pacbio-variant-version reflects the specification version that the file contents should adhere to.
The sequence-region headers describe the names and extents of the reference groups (i.e. reference contigs) that will be refered to in the file. The names are the same as the full FASTA header.
source-alignment-file and source-reference-file record absolute paths to the primary input files.
Feature lines
After the headers, each line in the file describes a genomic feature; in this file, all the features are potential variants flagged by the variant caller. The general format of a variant line is a 9-column (tab-delimited) record, where the first 8 columns correspond to fixed, predefined entities in the GFF standard, while the 9th column is a flexible semicolon-delimited list of mappings key=value.
The 8 predefined columns are as follows:
| Column Number | Name | Description |
Example
|
| 1 | seqId | The full FASTA header for the reference contig. |
lambda_NEB3011
|
| 2 | source | (unused; always populated with .) |
.
|
| 3 | type | the type of variant. One of insertion, deletion, or substitution. |
substitution
|
| 4 | start | 1-based start coordinate for the variant. |
200
|
| 5 | end | 1-based end coordinate for the variant. start<=end always obtains, regardless of strand. |
215
|
| 6 | score | unused; populated with . |
.
|
| 7 | strand | unused; populated with . |
.
|
| 8 | phase | unused; populated with . |
.
|
The attributes in the 9th (final) column are as follows:
|
Key |
Description |
Example value
|
|
coverage |
the read coverage of the variant site (not the variant itself) |
42
|
|
confidence |
the phred-scaled probability that the variant is real, rounded to the nearest integer and truncated at 93 |
37
|
|
reference |
the reference base or bases for the variant site. May be . to represent a zero-length substring (for insertion events) |
T, .
|
|
variantSeq |
the read base or bases corresponding to the variant. . encodes a zer-length string, as for a deletion. |
|
|
frequency |
the read coverage of the variant itself; for heterozygous variants, the frequency of both observed alleles. This is an optional field. |
|
The attributes may be present in any order.
The four types of variant we support are as follows. (Recall that the field separator is a tab, not a space.)
- 1.
-
Insertion.
Examples:
-
-
ref00001 . insertion 8 8 . . . reference=.;variantSeq=G;confidence=22;coverage=18;frequency=10 ref00001 . insertion 19 19 . . . reference=.;variantSeq=G/.;confidence=22;coverage=18;frequency=7/5
-
-
For insertions, start==end, and the insertion event is understood as taking place following the reference position start.
-
- 2.
-
Deletion.
Examples:
-
-
ref00001 . deletion 348 349 . . . reference=G;variantSeq=.;confidence=39;coverage=25;frequency=20 ref00001 . deletion 441 443 . . . reference=GG;variantSeq=GG/.;confidence=39;coverage=25;frequency=8/8
-
-
- 3.
-
Substitution.
Examples:
-
-
ref000001 . substitution 100 102 . . . reference=GGG;variantSeq=CCC;confidence=50;coverage=20;frequency=16 ref000001 . substitution 200 201 . . . reference=G;variantSeq=G/C;confidence=50;coverage=20;frequency=10/6
-
-
Compression
The gff metaformat is verbose, so for practical purposes we will gzip encode variants.gff files as variants.gff.gz. Consumers of the variant file should be able to read it in either form.