In the new tab, go to the NCBI website (https://www.ncbi.nlm.nih.gov/) and then click “BLAST” on the right-hand side. Notice that there are several ways of performing a BLAST search, and we will be covering two of them.
• While we won’t cover the reverse situation of performing a BLAST search when starting with a protein sequence, describe a potential scenario when you would have a protein sequence, but want to BLAST for a nucleotide sequence.
Click nucleotide BLAST (nucleotide -> nucleotide) button. This type of search expects a nucleotide reference sequence, and looks for similar nucleotide sequences in the database.
Input your sequence for the luxI gene.
Sometimes BLAST searches can return a lot of unwanted results. Since luxI is frequently used in plasmids, many cloning vectors are going to be listed, however for the context of this assignment we’re not interested in those. Under “Choose Search Set,” “Organism,” type in “taxid:29278” and from the list select “cloning vectors.” Then tick the “exclude” checkbox. This will remove them from the BLAST search.
Under “Optimize,” click “somewhat similar sequences.” Just ‘cuz.
• Before searching, what do you predict to be the main BLAST result? Why?
Click BLAST, and wait for results. Sometimes this can take several minutes, and certain times of the day during high traffic can also affect this.
• What is the best result (Species, E-value, coverage)?
• Are there any other interesting results? (You may have to scroll down a bit.) Write some here.
• Which of these would you expect? Why?
Scroll down past “Descriptions” and look at “Alignments.” This is where each individual result from the list in “Descriptions” is shown in a visual alignment. The top line is the reference gene used to query the database, while the bottom line is what it was found to match up with.