User GuidesSnapGene Pairwise and Multiple AlignmentCreate a Multiple Sequence Alignment

Create a Multiple Sequence Alignment

How do I create a sequence alignment in SnapGene?

SnapGene provides four third-party alignment tools that you can use to align three or more DNA and/or RNA sequences, or three or more protein sequences:

  • Clustal Omega
  • MAFFT
  • MUSCLE
  • T-Coffee

Click Tools → Align Sequences → Summary of Alignment Algorithms to learn about each algorithm.

Note that these algorithms do not check or automatically flip DNA or RNA sequences so that all are in the appropriate orientation. You should examine each sequence to confirm orientation, flip sequences if required, then perform the alignment.

Start the Alignment tool

To align DNA and /or RNA sequences, click Tools → Align Sequences → Align Multiple DNA Sequences.

To Align protein sequences, click Tools → Align Sequences → Align Multiple Protein Sequences.

Choose One or More Alignment Algorithm to Use

Use the check boxes to specify the algorithm/s you wish to use. You can choose multiple algorithms to create multiple alignments simultaneously if you wish.

SnapGene uses the published default settings for each alignment algorithm. For each algorithm you can click the Settings button to change advanced settings. Users should consult the algorithm websites, accessible via Tools → Align Sequences → Summary of Alignment Algorithms, to learn about advanced settings for each alignment algorithm.

Import Sequences for a New Alignment

You can choose sequences for alignment by a variety of methods:

Click Import Sequences to Align → Import Sequence Files to import sequence files from your computer.

Click Import Sequences to Align → Import Open Sequences to import sequences already open within SnapGene.

Click  Import Sequences to Align → Import Pasted Sequences to import sequences copied to the clipboard (Pasted data can be FASTA, FASTQ, GenBank/EMBL/DDJB, GCG, PIR/NBRF or PHYLIP formats).

Click Import Sequences to Align → Import NCBI Sequences to import sequences from NCBI using accession numbers.

You can also drag and drop one or more sequence files (including FASTA or GenBank multisequence files) into the alignment window.

Align the Sequences

Set one or Alignment algorithms to use.

Give the alignment a name.

Click Align to generate Alignments using the chosen algorithms.

The length of time required to generate an alignment will depend on:

  • The algorithm/s selected - use MAFFT or Clustal Omega for large or complex datasets
  • The size of the sequences (number of sequences and average length)
  • The complexity of the data set (relative similarity of all sequences)

View the Alignments

If multiple alignment algorithms were selected then use the "Aligned using" dropdown to select and view alignments created with each algorithm.

Configure the alignment view using the controls in the side panel.

If required, add a description of the alignment in the side panel.

To view features associated with a sequence click the disclosure triangle associated with the sequence. The amino acid sequence will also be shown for translated features (CDS).

To view features for all sequences, option-click (macOS) or ALT-click (Windows and Linux) on a disclosure triangle.

SnapGene 5.3 and later can display sequence features (if present) in alignments. See View Features in Multiple Sequence Alignments.