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106 Crabtree et al.17. Some clusters may contain adjacent or nearby genes from the same genome,because of tandem gene duplication. If the repeated genes are within the specified“neighborhood” distance then the system will automatically include only one copyof the relevant genomic subsequence in the comparative sequence display; thealternative, which is to include multiple copies of the same sequence that are offsetfrom one another, can be quite confusing visually.18. The order in which the genomes and/or sequences appear in the protein clusterdisplay may be set to a fixed default in Sybil, or one may click on a gene in thedisplay to force that genome and sequence to appear at the top of the image. Thisfeature was used in the TriTryp comparative annotation project (25) to designateone of the genomes as a reference against which the others were (manually)compared. Curators were able to traverse three genomes simultaneously by navigatingalong the fixed reference sequence using a modified version of the proteincluster display shown in Fig. 2.19. The purpose of this weight function is simple; by setting the edge weights in thisfashion one ensures that whenever geneA, geneB, and geneC are arranged fromtop to bottom in the cluster display, the algorithm will always prefer shortermatches (edges) to longer ones; geneA will be connected to geneB and geneB willbe connected to geneC, instead of drawing one long match between geneA andgeneC, followed by another between geneA and geneB, or geneB and geneC.20. The authors use Kruskal’s algorithm (26), as implemented by the Perl moduleGraph::Kruskal.21. This approach does not always produce an ideal figure layout, but in the authors’experience it does well in simple cases, and in complex cases it will at leastremove the redundant matches.22. The gene–gene matches are drawn differently depending on whether the genes inquestion appear in the same orientation. This provides an easy-to-see visual cuefor genes that are inverted in one genome relative to the others.23. In order for this to work a small patch must be made to Bio::Graphics::Panel, inwhich calls to GD::Image::colorAllocate() are replaced with identical calls toGD::Image::colorResolve(). This change allows all the panels in the image toshare the same GD color palette.24. Sybil also supports Scalable Vector Graphics (SVG) http://www.w3.org/graphics/SVG (27) output. SVG format images can be converted to PDF with the ApacheBatik package see http://www.apache.org/ and http://xmlgraphics.apache.org/batik/contributors.html (28), which provides an easy way to generate high-resolutionimages suitable for presentation or publication.AcknowledgmentsThe authors would like to thank all the Institute for Genomic Research(TIGR) faculty and staff who made suggestions and contributed feedback atevery stage of the algorithm and software development process. This work isfunded by the National Institute of Allergy and Infectious Diseases (NIH-NIAID-DMID-04-34).