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Finding Polymorphisms to Differentiate Species

 

anntrottpods.jpg FiPoDS takes two (possibly very large) FASTA files and searches one against the other sequence by sequence looking for local alignment hits that are top scoring but not identical. For each alignment, priming sites are identified to generate an (optionally nested) primer design for amplifying segments containing SNPs and indels. This is intended (and we use it) for trawling through genomic and/or RNA data sets of pairwise taxa in order to find points of diagnostic divergence.

 

Download

Save this file to your desktop, and unzip the contents (Macs will probably do this automatically). The FiPoDS folder within contains the FiPoDS script and a couple of associated files.

 

Installation

The script requires that your computer has Perl installed. The first line of the script indicates the location of Perl, and may need to be changed to suit your system. Macs should be OK, as this script was originally created to run on a Mac. If you use Windows and don't have Perl installed, you can install the 'industry standard' free Windows Perl ActivePerl, which we believe is robust and safe to install.

 

FiPoDS also makes use of NCBI BLAST to search the FASTA files. The Mac version of BLAST is included, but if you use Windows or Linux you will need to replace this. The easiest way to do this is to download the BLAST program from the NCBI FTP site, and install it in the folder called BLAST within the FiPoDS folder created above. Alternatively, if you already have BLAST installed, you could change the FiPoDS script to point to your BLAST location.

 

Parameters

FiPoDS searches two FASTA files against each other, so first it needs to know the names of your two files. These are the only mandatory parameters.

FiPoDS can carry out reciprocal BLAST searching between your two files (recommended), and needs to be told not to if you don't want it. This parameter is optional.

For the primer design stage, a target length in bp of the final PCR products is required, and also minimum and maximum lengths. Similarly, minimum and maximum GC content of products are required. These parameters are optional. If you do not specify any, the default values will generate primer designs suitable for amplification of even ancient or damaged DNA.

For further information in detail about parameters and how to change the defaults, see the script header in the file fipods_1.0.1.pl.

For quick start, all you need to do is edit the file fipods.ini, and put in the names of your FASTA files.

 

Running

On Macs and Linux, you can edit the file fipods.ini to add the names of your two FASTA files, then double-click the file FiPoDS.command. If you have installed the software in a place other than your desktop you can change the location in this command file.

FiPoDS is also intended to be run from the command line on any platform. You can put your chosen FASTA filenames and other parameters in the file fipods.ini or you can specify them explicitly on the command line. Explicit command line values override the defaults in fipods.ini. See the script header in the file fipods_1.0.1.pl for more information about command line options.

Output is generated to a comma-separated file with the name of your first FASTA file followed by .out. This file is easily explored in Excel.

 

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Licence

The software is provided free for academic use only. If you wish to use the software for another purpose please contact us. Full licence details under construction.

 

Citation

We have not yet published FiPoDs, so please cite the website if you use it in publication, or (FiPoDs Palmer, Moore and Allaby, University of Warwick, UK).

 

Please note that this is the first full release of this software, and we have not been able to test every possible combination of data sets, parameters, operating systems, and computing platforms. Please take this into account if using this software for publication purposes.

 

Changes since previous version

1.0.1 09/06/2008
Added fipods.ini functionality for optional supply of default values for parameters
Improved usefulness of comments

 

Known bugs

None.

 

 

The pod forms image is from a photograph of an original sculpture by Ann Trott and is used with the kind permission of the artist.