Section 3: Batch searches

So far, you have only performed a single sequence search. However, the Geneious BLAST interface also allows you to perform searches on multiple sequences at once.

To demonstrate batch searching, select the liver ESTs file and click BLAST. The search options should show how many sequences you have selected for the batch search. Because these are nucleotide sequences the default search algorithm chosen is Megablast. This program will find only high similarity matches so may return no hits if your sequences are from a less common species. Change this to blastn for a more sensitive search, and keep the other settings at their defaults. Your search dialog should look like this:


Clicking the Search button now will create a folder for the batch search. Be patient, it may take some time for the results of all searches to be returned. You can do other work with Geneious while the batch search is running.

When the results complete, they should look something like this


As with the single sequence search, each folder contains the results for the query sequence indicated by the name of the folder.


Query-centric searching

When batch BLASTing large numbers of sequences it can be impractical to have a separate search result folder created for each query. Instead, it may be more useful to only return a single alignment for each query. To do this, select the liver ESTs file again, click BLAST and under Results choose Query-centric alignments only. Change the Maximum Hits to 5 and click Search.

This time only a single results folder is returned containing an alignment for each query sequence.


The Search Hit annotation on each sequence contains the statistics for the blast match, such as E value, pairwise identity, sequence length etc. You can display this information in tabular format in the Annotations table. To do this, select all the alignments and switch to the Annotations tab. The default parameters are not set up for BLAST annotations, so we will change them by clicking the Columns button. In the Columns list, uncheck #Intervals, Direction, Maximum, Minimum, Name, Sequence and Type, and instead check %Pairwise Identity, E value, Hit start, hit end, Organism, Query, Query start, Query end and Sequence Name. You can arrange these columns as you wish by dragging and dropping them, and you should end up with a table that looks something like this:


This table can be exported in .csv format if required.


Filtering sequences using BLAST

When batch BLASTing there is also an option to bin into "has hit" vs "no hit" in database under Results. This is useful for filtering sequence reads for contamination from a particular non-target species such as human or E. coli. Instead of returning a sequence hit, the query sequences are sorted into "has hit" and "no hit" sequence lists.

You have now covered the basics of using BLAST. See Section 4 for more detailed information on the search options.

Section 4: Search options