Example 1: Single Annotation with Evidence (the simple life)

perl xGAEVAL.pl --clean_db --GFF examples/example1.gff3

Description:

This example illustrates the least complex and most often encountered usage of GAEVAL. In this case the example1.gff3 file encodes a single gene annotation ( the Arabidopsis thaliana locus At3g20090.1 ). In terms of the gff3 format this includes the gene, mRNA, exon, and CDS lines describing the structure and properties of this gene encoding transcript. Also included in example1.gff3 are match lines describing the alignments of three cDNA sequences (gi numbers: 42463391, 42460729, and 17381119) and two EST sequences (gi numbers: 19877306 and 59927596).

GAEVAL Results:

Annotation Summary

1: Annotation: At3g20090.1
2:  Genomic Source: 3
3:  Structure: join(7015803..7015913,7016352..7016407,7016733..7017561,7017643..7018366)
4:  Open Reading Frame: 7017052 to 7018293   
5:  5` UTR length:  484
6:  CDS length:     1159
7:  3` UTR length:  73
8:  Total length:   1716

This section is included in the basic report for each individual annotation and includes the follow information.

line 1: The geneId / name of the annotation
line 2: The id, gi, chromosome number, or name of the genomic region in which this annotation is located
line 3: The GenBank format structure description of this annotation
line 4: The open reading frame boundaries defined by the CDS of this annotation
line 5: The length of the 5` untranslated region as defined by the CDS
line 6: The length of the CDS (coding sequence)
line 7: The length of the 3` untranslated region as defined by the CDS
line 8: The total length of the annotated transcript

Structure Analysis

 1:  Integrity Score (0-1): 0.99
 2:  Exon Sequence Coverage: 100%
 3:  5` Terminus
 4:   Evidence supports the extension of this annotation boundary by 88 bases
 5:  3` Terminus
 6:   Evidence supports the extension of this annotation boundary by 2382 bases
 7:  Introns (total|confirmed|unsupported): 3 | 3 | 0
 8:  Individual Intron Support:
 9:   Intron 1 ( 7015914..7016351 )
10:    Supporting Alignments: 1
11:   Intron 2 ( 7016408..7016732 )
12:    Supporting Alignments: 1
13:   Intron 3 ( 7017562..7017642 )
14:    Supporting Alignments: 3

This section is also included in the basic report for each individual annotation and includes the follow information.

line 1: An integrity score relating the reliability / stability of the current annotation with respect to given evidence alignments
line 2: The percentage of the exonic sequence defined by the annotation which is supported by evidence alignments.
line 4: The support clues for the annotated transcription initiation site
line 6: The support clues for the annotated transcription cleavage / polyadenylation site
line 7: The count of introns (total, confirmed, and unconfirmed)
line 8+: Support for individual introns defined by the annotation. This includes the count of evidence alignments supporting each intron.

Incongruency Analysis:

 1: Incongruency Analysis:
 2:  No Ambiguously Overlapping Annotations Detected

 3:  Incongruent Introns Detected:
 4:   7016408..7016732
 5:     Conflicting Intron
 6:      Conflicting evidence: 1
 7:      Supporting evidence: 1
 8:   7018300..7018455
 9:     Alternative Intron
10:      Supporting evidence: 1
11:   7018469..7018493
12:     Additional Intron
13:      Supporting evidence: 1
14:   7018584..7018620
15:     Additional Intron
16:      Supporting evidence: 1
17:   7018701..7019102
18:     Additional Intron
19:      Supporting evidence: 1
20:   7019938..7020031
21:     Additional Intron
22:      Supporting evidence: 1

23:  No Complex Transcript Processing Detected

This section describes incongruencies found between the annotation and the supplied evidence alignments.

line 2: GAEVAL checked for erroneous overlap with adjacent annotations (if supplied) and found none. For more on this see example 2
line 3: Incongruent introns were detected
lines 4-7: Details about the incongruency found for the intron spanning genomic region 7016408..7016732
- line 5: The incongruence type is 'Conflicting Intron'
- line 6: The count of evidence alignments conflicting with this intron
- line 7: The count of evidence alignments supporting this intron
lines 8-22: More details about incongruent introns
- line 9: The incongruence type is 'Alternative Intron'
- line 12,15,18,21: The incongruence type is 'Additional Intron'
line 23: GAEVAL checked for complex annotation structure (Fusion, Fission, Alternative termination) and found none. For more on this see example 6

Discussion:

This example shows how GAEVAL can be used to quickly evaluate evidence for an annotation structure. For example, the At3g20090.1 locus in this scenerio is 100% covered by cDNA/EST alignments with all three of its annotated introns confirmed by at least 1 alignment. This along with the exceptable lengths of 484 bases and 73 bases for its 5` and 3` UTRs give this annotation and exceptional 0.99 integrity score. However, the GAEVAL report also details possible changes / additions applicable to the annotation. Such changes include the support for extending the 5` and 3` annotation boundaries and the documentation of possible alternatively spliced isoforms or additional introns.