Computerized STR Analysis

STR analysis is primarily a matter of testing for the presence or absence of products of particular sizes, representing the various alleles. This exercise uses analysis software to help you organize and manage data from PCR results.

Download and install Genographer.
Log into the PCR Simulator. You must have your own user name for this exercise. For forensics exercises, use "Template set: HUMAN", and check the box next to "Use named primers".
Run the reactions as usual. Be sure your gel looks reasonable.
Go back to the PCR setup page. At the bottom of the paeg, click "other formats". This will take you to a new page where you can retrieve the results of the PCRs you just ran as electropherograms.
- Choose a molecular size standard (such as "ladder-100bp")
- Leave the "archive method" set to "zip" (unless you really want a tar.gz file.)
- Click "get chromatograms"
- Click the "get size standards file" button to download a copy of the "standards.cfg" file containing the sizes of the marker bands. Replace the file of the same name in the Genographer directory.
Extract your set of chromatograms from the "zip" archive.
- On WindowsXP, right-click the "zip" file and select "Extract here".
- In MacOSX, double click it.
Launch Genographer (probably by double-clicking the genographer.class file - if all else fails, read the directions.)
Choose "File: Import". Navigate to the directory with your extracted SCF files.
Click "Add all" to use the whole set of SCF files. Be sure to select "Files of type: SCF". Click "Import".
In the SCF Parameters dialog box, choose the following:
- Data Channel = Blue (Channel 3)
- Standards Chan. = Red (Channel 1)
- Size Method: Local Southern
- Size Standard: ladder-100bp (or whatever standard you chose when retreiving your PCR results as SCF files)
- Min Peak Height: 1.0
- Min Peak Width: 1.0
You should now be able to point to a band in Genographer and read its size.
The SCF files only contain data up to 1000 bp, so you can only use them for analyzing bands smaller than that limit.

This is an example of a set of PCR reactions loaded into Genographer. These reactions were run under the username "scftest", so you can get a copy by using that user name on the PCR trace download page. PCRs were run using the pairs of named primers against the H_sapiens reference genome with the standard conditions.

Ethidium gel	Genographer "gel"

Pointing to a band in Genographer "gel" display will display it's size on the status line.