The final annotation file was generated after gene-ID mapping, GO term assignment, annotation augmentation, and generic GO-slim processes.
The first objective is the assignment (annotation) of Roche 454 reads of a single genotype to three categories: (1) characterized gene reads, (2) characterized repeats, and (3) uncharacterized reads.
The GO terms associated with each BLAST hit were retrieved (mapping step) and GO annotation assignment (annotation step) to the query sequences was carried out using the following annotation score parameters; E-Value Hit Filter (default=1.0E-6), Annotation Cut-Off (default =55), GO-Weight (default=5), Hsp-Hit Coverage Cut Off (default = 0).
Following automatic assignments, annotation of the identified ORFs was performed based on sequence similarity searches against sequence databases and subsequent manual curation and annotation using GenDB 2.4 [ 28].
In all cases, longer sequence reads resulted in more sensitive assignment of annotations to the DNA sequences, as seen before (Wommack et al., 2008).
*For At. thiooxidans interactions were assigned based on ortholog assignment Interaction annotation data (Table 1B) were sourced from STRING [ 40], MetaCyc [ 41] and KEGG [ 42].
Using a conservative approach this nomenclature system is applied during manual annotation, where the specificity of the gene name reflects the confidence in the assignment and annotation is consistent with the best available evidence.
In alternative, raw reads can be also used for direct assignment and annotation, though their short length may limit the exhaustivity of the results.
If the anomalous pump correlated with an incorrect peak assignment the annotation was corrected using the editing features of AutoEPG.
Using the proportion of DNA and mRNA sequences that had homology to known proteins, we were able to make phylum-level taxonomic assignments using annotations from the SEED databases [18] (Fig. 2).
