To make a request to a local BLAST database do the following:

• If you’re using BLAST open Tools ‣ BLAST ‣ BLAST Search.
• If you’re using BLAST+ open Open Tools ‣ BLAST ‣ BLAST+ Search.

If there is a sequence opened you can also initiate the request to a local BLAST database from the Sequence View:

• If you’re using BLAST select the Analyze ‣ Query with BLAST item in the context menu or in the Actions main menu.
• If you’re using BLAST+ select the Analyze ‣ Query with BLAST+ item in the context menu or in the Actions main menu.

The Request to local BLAST database dialog will appear:

<center>
     <br>
     <img src="/wiki/download/attachments/14059181/Making Request to Database.png"/>
     <br>
</center>

The following general options are available:

Select search - here you should select the tool you would like to use. If the query sequence is a nucleotide sequence then blastn, blastx and tblastx items are available. For a protein sequence the items are blastp and tblastn.

Expectation value - this option specifies the statistical significance threshold for reporting matches against database sequences. Lower expect thresholds are more stringent, leading to fewer chance matches being reported.

Culling limit - the maximum number of hits that will be shown (not equal to number of annotations). The maximum availablle number is 5000.

Search for short, nearly exact matches - automatically adjusts the word size and other parameters to improve results for short queries.

Megablast - select this option to compare query with closely related sequences. It works best if the target percent identity is 95% or more, but it is very fast.

Database path - path to the database files.

Base name for BLAST DB files - base name for the BLAST database files.

You can see the description of the annotation saving parameters here.

The following advanced parameters are available:

<center>
     <br>
     <img src="/wiki/download/attachments/14059181/Making Request to Database_1.png"/>
     <br>
</center>

Word size - the size of the subsequence parameter for the initiated search.

Gap costs - costs to create and extend a gap in an alignment. Increasing the Gap costs will result in alignments which decrease the number of Gaps introduced.

Match scores - reward and penalty for matching and mismatching bases.

Filters - filters for regions of low compositional complexity and repeat elements of the human’s genome.

Masks for lookup table only — this option masks only for purposes of constructing the lookup table used by BLAST so that no hits are found based upon low-complexity sequence or repeats (if repeat filter is checked).

Mask lower case letters — with this option selected you can cut and paste a FASTA sequence in upper case characters and denote areas you would like filtered with lower case.

The following extension options are available:

<center>
     <br>
     <img src="/wiki/download/attachments/14059181/Making Request to Database_2.png"/>
     <br>
</center>

For gapped alignment - X dropoff value (in bits) for gapped alignment.

For ungapped alignment - X dropoff value (in bits) for ungapped alignment.

For final gapped alignment - X dropoff value (in bits) for final gapped alignment.

Multiple hits window size - multiple hits window size.

Perform gapped alignment - performs gapped alignment.