The Specific Primer & Amplicon Design Software (SPADS) has been specifically developed for (1) the selection of specific regions within genes and (2) the design of primer pairs picked to amplify such regions.
SPADS is described in:Thareau V., Déhais P., Serizet C., Hilson P., Rouzé P. and S. Aubourg (2003) Automatic design of gene-specific sequence tags for genome-wide functional studies. Bioinformatics 19, 2191-2198.
The CATMA GSTs are 150 to 500 bp genomic fragments amplified by PCR, a maximum of 50% of which may be intron sequence. Most have been selected to share no more than 70% identity with any other sequence in the Arabidopsis genome. However, because of the inherent duplicated nature of the Arabidopsis genome, this will not be possible for all genes. Lower-specificity GSTs will be produced for the remainder, and they will be flagged as such in the database.
All CATMA GST PCR amplicons are examined by agarose gel electrophoresis, and are only accepted if a single clear band of the correct size is observed (example gel image). Where possible, GSTs are amplified from BAC templates. Over 1,400 of the 24,576 GSTs have been sequence verified, and no incorrectly amplified sequences have been found.
The CATMA GSTs are flanked by a limited set of sequences allowing for PCR reamplification and preventing well-to-well cross-contaminations which often plague the storage and dissemination of large-scale clone collections. Full sequence information and design parameter for these extensions is available here.