BIT150 – Lab3
Multiple sequence alignment and Phylogenetics
Copy 08_Lab3 from Z: to C:, and open the file ‘FT proteins for MEGA.doc’.
Objective: Perform multiple sequence alignments, calculate distance matrices, and
construct phylogenetic trees, to understand and interpret relationships between species.
Activities:
A. Creating Multiple Sequence Alignments (MSA)
In this example, we will create a multiple alignment of protein sequences that will be
imported into the alignment editor using different methods. Multiple protein sequence
alignment is a central tool to infer protein function, predict protein secondary structure,
and identify residues important for protein specificity.
A1. Start MEGA4 by using Start\Programs\BioInformatics\MEGA4.
A2. In the MEGA4 window, go to Alignment|Alignment Explorer/CLUSTAL. Select
‘Create a new alignment’, and click on OK. Click on [NO] for protein sequence
alignment.
A3. Sequences can be entered either from FASTA files or by hand. We will enter the
sequences by hand, one by one. In the Alignment Explorer window, go to Edit|
Insert Blank Sequence or click on , and repeat it to generate 8 blank
sequences. Right-click on the blank sequence name and edit the sequence name
for each protein sequence, as it is in the Word document ‘FT Proteins for MEGA’.
Copy and paste each sequence.
A4. Go to Edit|Select All to select every site for all the protein sequences in the
alignment.
A5. Go to Alignment|Align by ClustalW or click on to align the selected protein
sequences using the ClustalW algorithm.
A6. Save the current alignment by selecting the Data|Save Session. Save it as ‘FT.mas’.
This will allow the current alignment to be restored for future editing. Also,
export it (Data|Export Alignment|FASTA format) as both a FASTA file
(‘FT.fas’) and a MEGA file (‘FT.meg’).
B. Generating a publishable MSA using BoxShade
1
