We are going to use some real data from a sequencing project to investigate many of the functions of sequencher.
The sequence files are available either Unix tarred gzip'd archive (should work with Macs and PCs too). For PCs you will need the application WinZip that you can find on this site: VersionTracker. The sequences are also available as a WinZip self extracting archive, or as raw data (be sure to download them as binary files). To download these files on a mac click the mouse button and hold until a menu appears. Choose "Save link as". On a PC, right click on the link and choose "Save link as".
Open the folder that contains the sequences, there should be 11 sequences, numbered 1-11. If you do not have the right number of sequences, or are having problems with the downloading or extraction email Rob Edwards
Once you have downloaded and extracted the files and started Sequencher we are ready to begin. Highlight all the sequences in the folder and drag them onto the sequencher window. Sequencer will read all the trace files and you will see something like that shown below.
In the first column there is an icon 
that indicates that the item is a sequence. We will see this change after we have assembled sequence. The second column contains the file or contig names. At the moment it only has the file names. The third column contains the size of the sequence in bp. The next two columns have a brief description about how the sequence was generated. This information is gleaned from the sequence file. Finally, there are the date, time, and comments about the sequence. We will come back to the comments later.
At the top of the window there are five buttons. The up arrow will remove these buttons. The next three buttons deal with the assembly, and we will return to these. Finally there is a trash can icon to delete one or more sequences.
To begin with we will take a look at one sequence. Click on the first sequence, sequence01.ab1. You will see a window like this appear:
This window shows the sequence generated from the chromatogram. This particular sequence is 851 bp long. You will note that most of the sequence is A,G,C, or T, but there are several N's in the sequence, particularly near the end of the sequence. We will come back to these N's later in the tutorial
At the top of this window you see eight buttons. The first button Overview (try it!) gives an overview of the sequence, where the start and stop codons are, and so on. You may have to alter the Options to see all the features. The second button Cut Map gives a restricition map of the sequence like this:
You can change the enzymes that are listed by clicking on the Select Enzymes button. You can also change the look by clicking on the Options to see some of these things. See if you can figure out what filter I used for these restriction enzymes. To return to the sequence view and continue, click on Bases.
In the bases view, the next two buttons: 
and 
bring the start and stop codons for each frame and the restriction digest into view in this window.
The Ruler button provides a ruler with a number of different functions as shown below:
There are 15 buttons on the ruler:
If you incorporate many of these changes the standard nucleotide sequence view you can end up with something looking like this:
The final button here Show chromatogram is arguably the most important and that will be dealt with in section 2 of the tutorial.
