Supplementary Method S1: Alignment of physical and genetic maps
A multiple pass technique was used to generate putative associations of sequence scaffolds to a genomic mapping. Information from all available sources was used to generate a series of increasingly refined mappings: fingerprint gels and FPC assemblies1 were generated as per Zhu et.al.2 and incorporated into the MagnaportheDB database 3, Arachne 4 was used to generate associations of assembly scaffolds, sequence contigs, and BAC end reads; genetic maps for the ancestors of strain 70-15 were obtained from Nitta et al.5. A conservative approach was adopted where in only scaffolds unambiguously associated with a chromosome were assigned chromosomal locations. Order and orientation of scaffolds were determined by locating two or more markers with the genetic map. Scaffolds whose location could not be unambiguously determined were placed in an artificial chromosome named zero (0). Description of each pass is as follows:
Pass 1: BAC library end sequences were incorporated into the whole genome assembly process via the Arachne 4 sequence assembler. Two explicit datasets were extracted from the Arache assembler results. One associatingBAC to assembly contigs [BAC-CONTIG] and the other associating scaffolds to contigs [SUPERCONTIG].
Pass 2: Associations of assembly scaffolds to the genetic map were generated based on physical mapping information fromMagnaportheDB 3 and the SUPERCONTIG data set. The association was scaffold -> sequence contig -> BAC end sequence -> fingerprint contig -> marker -> chromosome. A noise reduction filter was applied by identifying maxima of these associations. Associations that did not have significant maxima were marked for additional work.
Pass 3: Fingerprint contigs responsible for generating indications of contradictory scaffold to chromosome associations where reviewed. Those contigs based on a relatively weak relationship between strongly associated groupings of BACs were split into multiple contigs. Any newly unambiguous associations were added to the result set.
Pass 4: A composite genetic map [GCOMPOSIT] was formed based on genetic maps for the ancestors of strain 70-15 5. Markers from the same chromosome of all strains were combined together and a relative ordering was established for those markers on the chromosome. The association scaffold -> sequence contig -> BAC end sequence -> marker -> chromosome of scaffolds to chromosomes was created based on theSUPERCONTIG, BAC-CONTIG, andGCOMPOSIT datasets. The new associations were merged into the previous results. Ambiguous associations where identified and flagged for additional work.
Pass 5: The set of scaffold to chromosome associationsflagged as being contradictory were subject to expert review. The evidence for the association was examined to identify impossible relationships and those resulting from repetitive sequence. These relationships were eliminated and any newly unambiguous associations were added to the result set.
Pass 6: For the remaining unassigned scaffolds whose size exceeds that of a BAC, two or more specific genetic markers were identified. The sequence of these markers was obtained and located within the sequence contigs viasimilarity search using the blast program6. The associationscaffold -> sequence contig ->marker -> chromosome of scaffolds to chromosomes was created based on theSUPERCONTIG andGCOMPOSIT datasets. Any newly unambiguousassociations were merged into the previous results.
Out of 187 genetic markers, 133 were placed on the BAC physical map by hybridization. These BACs were placed on the assembly through their end sequences. In total, 126 markers were anchored on the genome assembly.
1.Soderlund, C., Humphrey, S., Dunham, A. & French, L. Contigs built with fingerprints, markers and FPC v4.7. Genome Res.10, 1772-87 (2000).
On-line:
2.Zhu, H., Blackmon, B. P., Sasinowski, M. & Dean, R. A. Physical Map and Organization of Chromosome 7 in the Rice Blast Fungus, Magnaporthe grisea. Genome Res.9, 739-750 (1999).
3.Martin, S. L. et al. MagnaportheDB: a federated solution for integrating physical and genetic map data with BAC end derived sequences for the rice blast fungus Magnaporthe grisea. Nucleic Acids Res. 30, 121-124 (2002).
On -line:
4.Batzoglou, S. et al. ARACHNE: A Whole-Genome Shotgun Assembler. Genome Res.12, 177-189 (2002).
5.Nitta, N., Farman, M. L. & Leong, S. A. Genome organization of Magnaporthe grisea: intergration of genetic maps, clustering of transposable elements and identification of genome duplications and rearrangements. Theor. Appl. Genet.95, 20-32 (1997).
6.Altschul, S. F., Gish, W., Miller, W., Myers, E. W. & Lipman, D. J. Basic local alignment search tool. J. Mol. Biol.215, 403-410 (1990).