BLAST

Jump to Section:

• Accessing BLAST Feature
• Running BLAST
• Downloading Sequences

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Overview

BLAST (Basic Local Alignment Search Tool) is a suit of programs hosted by NCBI that find regions of similarity between sequences by aligning query sequences against a database of known sequences. BLAST can be used to confirm taxa identified by CZ ID and annotate sequences. There are multiple types of BLAST (click here for details); however, CZ ID currently only supports BLASTN and BLASTX. Here we describe how to run BLAST from CZ ID's Sample Report page. Note that you can also download sequences and run BLAST through NCBI's website.

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Accessing BLAST Feature

You may want to use BLAST to confirm the detection of a taxon of interest. The BLAST feature  can be accessed from two locations in the Sample Report page, namely the Sample Report Table (taxon row) and the Coverage Visualization Panel. 

Accessing BLAST from the Taxon Row in the Sample Report Table

To access the BLAST feature directly from the Sample Report Table:

1. Hover over the taxon name of interest to see analysis icons.

2. Click the BLAST icon ().

Accessing BLAST from the Coverage Visualization Panel

To access the BLAST feature from the Coverage Visualization Panel:

1. Hover over the taxon name of interest to see analysis icons.

2. Click the Coverage Visualization icon (). This will open the Coverage Visualization Panel at the bottom of the page.

3. Click the BLAST icon () found in the right-hand side of the Coverage Visualization Panel.

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Running BLAST

To run BLAST:

1. Click the BLAST icon through the Sample Report Table or Coverage Visualization Panel (see Accessing BLAST Feature for details).

2. A modal will appear for you to select the BLAST type you wish to run. Click "Continue" after making your selection.

BLASTN vs BLASTX

BLASTN is the standard BLAST program that searches the nucleotide database (NT) using a nucleotide query sequence, whereas BLASTX is used to search the protein database (NR). BLASTN is an integral part of the metagenomic analysis workflow that can be used to:

• • Confirm taxon matches.
  • Gather relevant alignment metrics for publication (e.g., E-value, % identity, and bit score).  
  • Find contextual data (related sequences represented in the database) for phylogenetic trees .
  • Perform quality control for contigs generated through SPAdes (de novo assembler). By "blasting" contigs, users can identify chimeras (contigs formed by two or more reads that have been incorrectly assembled together) and determine contig orientation for downstream analyses.

BLASTX translates nucleotide query sequences and compares translated amino acid sequences to the protein database.  It can be used for:

• • Identifying coding regions and encoded proteins
  • Confirming novel viral sequences that only have matches in the NR database

3. Once the type of BLAST is specified, another modal will appear for you to select which sequences you would like to run. To select sequences, check the boxes for contigs or reads of interest. Click Continue after selecting the sequences for BLAST.

BLASTN Modal: CZ ID will allow up to 3 of the longest contigs that aligned to the NT database to run BLASTN. If no contigs aligned to the NT database, CZ ID will send up to  5 reads that aligned to the NT database. The modal will automatically show the longest contigs or randomly selected reads available for the analysis.

BLASTX Modal: CZ ID will allow up to 3 of the longest contigs that aligned to the NT or NR database to run BLASTX. Note the NT hits (default) and NR hits tabs for you to select which sequences you would like to run. If no contigs aligned to the NT database, CZ ID will send up to 5 reads that aligned to the NT database. The same logic applies for NR hits if you choose to select sequences aligning to NR. The modal will automatically show the longest contigs or randomly selected reads available for the analysis.

4. A message will appear indicating that you will be redirected to NCBI's BLAST service. Click Continue to send your sequences to NCBI.  

5. An NCBI BLAST page will appear with default parameters. To run BLAST, simply click “View       report”.  

6. The BLAST Results page will appear after the analysis is done. If you ran multiple sequences, use the "Results for" dropdown menu to view the BLAST results for other sequences included in the analysis. Read about BLAST result interpretation in our Guide to BLAST or see NCBI's guide describing the BLAST Results Page.

7. Once you evaluate your BLAST results, you can annotate whether the taxon was confirmed ("Hit"), "Not a hit", or "Inconclusive" using the annotation feature. 

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Downloading Sequences for BLAST 

If you are having trouble running BLAST from CZ ID or you would like to run BLAST using additional contigs or reads that are not listed in the BLAST modal, you can download sequences and submit sequences directly through NCBI's BLAST website. 

To download sequences:

1. Hover over the taxon row and click the Download icon.

2. Select if you would like to download contigs or reads from the dropdown menu.

3. Go to NCBI's BLAST website, select the BLAST type of interest, and submit the contigs and/or read sequences of your choice. Read about BLAST types and how to interpret results here.