Aligning Sequences

Once you acquire sequences (which I covered in my previous blog post), you must then align the sequences before constructing the phylogenetic tree. In MEGA, Two alignment methods are provided: ClustalW and MUSCLE. Either can be used, but in general MUSCLE is preferable. In the Alignment menu in MEGA, I chose the MUSCLE button, and then I clicked on the "Align Codons" button. There were two choices: Align DNA and Align Codons, and since my sequence was a DNA coding sequence I chose Align Codons, which ensure that the sequences are aligned by codons, a much more realistic approach than direct alignment of the DNA sequences because that avoids introducing gaps into positions that would result in frame shifts in the real sequences. Once I started the alignment process in the program, it took about two minutes. Next, I exported my file in the correct format so I would be able to use it to estimate my phylogenetic tree later. 

Here is my final aligned file:

Once I completed the alignment, I noticed that gaps were introduced into the sequences. Those gaps represent historical insertions or deletions, and their purpose is to bring homologous sites into alignment in the same column. Justas a phylogenetic tree is an “estimate” of relationships among sequences, an alignment is just an estimate of the positions of historical insertions and deletions.