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Next , 32predictions ranked below the top 1% from VLS were randomly selected and tested experimentally on 1000001 and 1000006 to demonstrate that the obtained hit rate for the top 1% was appreciably better than the background.
The B2G program uses BLAST to find homologous sequences to input sequences and extracts GO terms (Gene Ontology) to each obtained hit using existing annotations.
Annotation of ESTs and extraction of GO terms to each obtained hit using existing annotations was done using the BLAST2GO v1 program [ 31] as previously described [ 27].
Finally, to identify other P. salinus MITE-related sequences in the A. castellanii genome, we used every MITE sequence identified in P. salinus as queries for homology searches (BLASTn) against the locally installed most recent assembly of the A. castellanii genome (Acas_2.0), and we manually checked every obtained hit.
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GO terms were derived from the obtained hits and for each sequence they were merged with the already assigned terms.
Some sequences obtained hits to multiple active site domains.
Blast2GO retrieves the most significant GO terms associated with the obtained hits to the query sequence.
For all obtained hits, t-tests were calculated to validate a statistical significance at α = 0.1.
All obtained hits were further assessed using Pfam domain analysis and reciprocal Blast search as described earlier.
We validated all the obtained hits by mapping functional domains and active site residues, yet the possibility of obtaining false positives cannot be ignored.
The title and abstract of the obtained hits were evaluated and articles referring to the gross morphology were obtained and assessed in detail.
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Justyna Jupowicz-Kozak
CEO of Professional Science Editing for Scientists @ prosciediting.com