Creating Multiple Sequence Alignments

In this example, we will create an alignment from protein sequence data that will be imported into the alignment editor using different methods.

Ex 1.0.1: Start MEGA by double-clicking on the MEGA desktop icon, or by using the Windows start-menu to click on the MEGA icon located in the programs folder.

Ex 1.0.2: Launch the Alignment Explorer by selecting the Alignment|Alignment Explorer/CLUSTAL menu command.

In order to align sequences contained in a Sequence Data File, do the following:

Ex 1.1.1: Add unaligned sequences from the hsp20.fas example file into the Alignment Explorer by clicking selecting the Data|Open|Retrieve Sequences from File menu command.

Ex 1.1.2: Select the Edit|Select All menu command to select every site for all sequences in the alignment.

Ex 1.1.3: Select the Alignment|Align by ClustalW menu command to align the selected sequences data using the ClustalW algorithm.

Ex 1.1.4: Save the current alignment session by selecting the Data|Save Session menu item. This will allow the current alignment session to be restored for future editing.

Ex 1.1.5: Exit the Alignment Explorer by selecting the Data|Exit Alignment Explorer menu item. A message will appear asking if you would like to save the data to a MEGA file. Choose �YES,� and then a �Save As� dialog box will appear. Enter hsp20_aligned.meg as the file name, and click the �Save� button. An input box will appear asking for a title for the data. Enter �HSP 20 Aligned by MEGA� as the title, and click the �OK� button. Another dialog box will appear asking you if the sequence data is protein coding. In this case, click �Yes.� A final dialog box will appear asking you if you would like to open the data file in MEGA. Click �Yes.�

Ex 1.2.1: If the Alignment Explorer already contains sequence data, select the Alignment | Create a new alignment menu command to create a new alignment. Choose �YES� on the dialog box that appears to indicate that you are creating a DNA sequence.

Ex 1.2.2: Activate the Web Explorer tab by selecting Web|Query Gene Banks from the menu.

Ex 1.2.3: When the NCBI Entrez site is loaded, select either the nucleotide or protein database, enter a search term into the search box, and press the �GO� button.

Ex 1.2.4: When the search results are displayed, select the specific search item and choose �Sequence� from the menu bar. Press the �Add to Alignment� button located to the left of the address box. This will display the Web Fetch dialog window.

Ex 1.2.5: Click the box to the left of each accession number whose sequences� information you would like to fetch from the web. When you are done, select accessions by pressing the �Fetch� button.

Ex 1.2.6: When the status column indicates that all sequences are fetched, press the �Send to Alignment� button to send the fetched sequence data to the Alignment Explorer.

Ex 1.2.7: Align the fetched data using the steps detailed in Ex 1.1.2 � Ex 1.1.5.

You may also open a trace file in the Trace Data Viewer/Editor and send it directly to the Alignment Explorer.