The Basic Local Alignment Search Tool (BLAST) finds regions of local similarity between sequences, which can be used to infer functional and evolutionary relationships between sequences as well as help identify members of gene families.
Select the Blast tab of the toolbar to run a sequence similarity search with the BLAST (Basic Local Alignment Search Tool) program:
- Enter either a protein or nucleotide sequence (raw sequence or fasta format) or a UniProt identifier into the form field.
- Click the Blast button.
The following kinds of UniProt identifiers are supported:
|P00750-2||UniProtKB entry isoform sequence|
|P00750[1-20]||Part of UniProtKB entry sequence, from its 1st to 20th amino acid residue (inclusive)|
|A4_HUMAN||UniProtKB entry name|
If you select the Blast tab of the toolbar from a UniProtKB, UniRef or UniParc entry page, the current sequence is prefilled in the form. Jobs have unique identifiers, which (depending on the job type) can be used in queries (e.g. to get the intersection of two sequence similarity searches). Job identifiers and the related data are kept for 7 days, and are then deleted.
|Database||Database against which the search is performed: UniProtKB or clusters of sequences with 100%, 90% or 50% identity.|
|Threshold||The expectation value (E) threshold is a statistical measure of the number of expected matches in a random database. The lower the e-value, the more likely the match is to be significant. E-values between 0.1 and 10 are generally dubious, and over 10 are unlikely to have biological significance. In all cases, those matches need to be verified manually. You may need to increase the E threshold if you have a very short query sequence, to detect very weak similarities, or similarities in a short region, or if your sequence has a low complexity region and you use the “filter” option|
|Matrix||The matrix assigns a probability score for each position in an alignment. The BLOSUM matrix assigns a probability score for each position in an alignment that is based on the frequency with which that substitution is known to occur among consensus blocks within related proteins. BLOSUM62 is among the best of the available matrices for detecting weak protein similarities. The PAM set of matrices is also available. If “Auto” is set, the matrix will be selected depending on the query sequence length.|
|Filtering||Low-complexity regions (e.g. stretches of cysteine in Q03751, or hydrophobic regions in membrane proteins) tend to produce spurious, insignificant matches with sequences in the database which have the same kind of low-complexity regions, but are unrelated biologically. If “Filter low complexity regions” is selected, the query sequence will be run through the program SEG, and all amino acids in low-complexity regions will be replaced by X’s.|
|Gapped||This will allow gaps to be introduced in the sequences when the comparison is done.|
|Hits||Limits the number of returned alignments.|
Performing a BLAST search;
- Select the ‘Blast’ tab of the toolbar at the top of the page to run a sequence similarity search with the Blast program.
- Enter either a protein or nucleotide sequence or a UniProt identifier into the form field (Figure 37).
- Click the ‘Run Blast’ button.
- The Blast input page.
- Blast searches can be run directly from the ‘Blast’ button in UniProt entry pages. All relevant results pages (such as UniProtKB, UniRef, UniParc and tool results) allow you to run a Blast search.
The Blast result page shows an overview of the results, colour coded by sequence identity in descending order by score (Figure 38). You can re-order the results by E-Value, Score or Identity.
Filters and Views
You can use filters on the left hand side to narrow down your search results, e.g. to limit the results to a particular species. You can also map the results to UniProt protein databases UniProtKB, UniRef and UniParc. You can view results by taxonomy or in plain text format.
More detail about each result can be seen in the ‘Alignments’ table under the ‘Overview’ section, showing the query sequence aligned to each subject sequence.
You can view each alignment in more detail by clicking on the graphic or on the ‘view alignment’ link. You can also see information about E-Value, Score and Identity. To add more information, click on the ‘edit columns’ button. You can run another Blast search from here by selecting an entry using the checkboxes and clicking on the ‘Blast’ button. You can also align your Blast results by selecting them using checkboxes and then clicking on the ‘Align’ button. The results can be downloaded in various formats or be added to your Basket for later.