---
title: "annot - Gene Ontology Annotation with Blast2GO"
canonical: "https://help.biobam.com/space/DOC/792836/annot%20-%20Gene%20Ontology%20Annotation%20with%20Blast2GO"
format: markdown
---
> Macro (toc)

## Blast2GO Annotation Rule

<span style="color: #222222">This is the process of selecting GO terms from the GO pool obtained by the Mapping step and assigning them to the query sequences. In the current Blast2GO version this is the core type of functional annotation.</span>

GO annotation is carried out by applying an annotation rule (AR) on the found ontology terms. The rule seeks to find the most specific annotations with a certain level of reliability. This process is adjustable in specificity and stringency.

For each candidate GO an annotation score (AS) is computed. The AS is composed of two additive terms.

The first, direct term (DT), represents the highest hit similarity of this GO weighted by a factor corresponding to its EC.

The second term (AT) of the AS provides the possibility of abstraction. This is defined as an annotation to a parent node when several child nodes are present in the GO candidate collection. This term multiplies the number of total GOs unified at the node by a user-defined GO weight factor that controls the possibility and strength of abstraction. When GO weight is set to 0, no abstraction is done.

Finally, the AR selects the lowest term per branch that lies over a user-defined threshold. DT, AT and the AR terms are defined as given in Figure 1.

> Macro (anchor)



![image](media://ca55c67a-6858-4f80-a8e8-65d3a8a1aeaf)

**Figure 1:** Blast2GO Annotation Rule

  


To better understand how the annotation score works, the following reasoning can be done: When EC-weight is set to 1 for all ECs (no EC influence) and GO-weight equals zero (no abstraction), then the annotation score equals the maximum similarity value of the hits that have that GO term and the sequence will be annotated with that GO term if that score is above the given threshold provided. The situation when EC-weights are lower than 1 means that higher similarities are required to reach the threshold. If the GO-weight is different to 0 this means that the possibility is enabled that a parent node will reach the threshold while its various children nodes would not.

The annotation rule provides a general framework for annotation. The actual way annotation occurs depends on how the different parameters at the AS are set. These can be adjusted in the Annotation Configuration Dialog (Figure 2) and in the Evidence Code Weight Configuration Dialog (Figure 3).

  


  


|  |  |
| --- | --- |
| ![ecweight1.png](media://f8b392a0-18f1-4264-9ed3-f9b2ff26e58f) | ![2.png](media://b2714224-0d82-4167-be13-74b8078c9395) |

**Figure 3:** Evidence Code weight configuration

  


Successful annotation for each query sequence will result in a color change for that sequence from light-green to **blue** at the Main Sequence Table, and only the annotated GOs will remain in the GO IDs column.

  


## Annotate GOs from Blast Descriptions

<span style="color: #222222">This tool looks at every significant alignment (</span><span style="color: #222222">**Right-Click > Show Blast Result**</span><span style="color: #222222"> on a sequence) for each sequence and searches their description lines for GO ids. These GOs are now directly annotated to the sequence if the alignments similarity passes the desired minimum. Validation can also be applied and is recommended, it will remove intermediate GO terms.</span>

  


## Load Annotation Results (.annot)

<span style="color: #222222">Already made or existent annotation can be imported using the .annot format. For import purposes only, the .annot format allows also multiple annotations of the same sequence to be given in one single row, separated by commas, as shown above (Schema: Seq-Name <tab>GO(s) or EC(s) <tab>Sequence description):</span>

Blast2GO Annotation File (.annot):

<sup>Seq1 GO:0001234 glycolipid transfer protein-like</sup>  
<sup>Seq1 GO:0001264,GO:0004567,...</sup>  
<sup>Seq1 GO:0034567</sup>  
<sup>Seq1 EC:2.1.2.10</sup>  
<sup>Seq2 GO:0001234,... sorbitol transporter</sup>  
<sup>Seq2 GO:0001244</sup>  
<sup>Seq3 GO:0001234,GO:0004567,GO:0009123</sup>  
<sup>Seq3 EC:1.2.4.1, EC:3.1</sup>  
<sup>....</sup>

  
There are still other annotation functions available in the sub menu:

## Other Annotation Functions

- Remove Annotation. Delete Annotation results for the selected sequences.
- Filter Annotation by GO Taxa
- Validate Annotations. Blast2GO annotation generates lowest node annotations. This is not always guaranteed when Annotations have been imported or changed manually. This function can be run to ensure that no parent-child redundancy is present in the annotated set.
- Remove 1. Level Annotations
- Annotate GOs from Blast Descriptions allows to transfer GOs from the Blast hit descriptions to their sequences.