title: Gene discovery in EST sequences from the wheat leaf rust fungus Puccinia triticina sexual spores, asexual spores and haustoria, compared to other rust and corn smut fungi.
abstract: BACKGROUND
  Rust fungi are biotrophic basidiomycete plant pathogens that cause major diseases on plants and trees world-wide, affecting agriculture and forestry. Their biotrophic nature precludes many established molecular genetic manipulations and lines of research. The generation of genomic resources for these microbes is leading to novel insights into biology such as interactions with the hosts and guiding directions for breakthrough research in plant pathology.
  
  
  RESULTS
  To support gene discovery and gene model verification in the genome of the wheat leaf rust fungus, Puccinia triticina (Pt), we have generated Expressed Sequence Tags (ESTs) by sampling several life cycle stages. We focused on several spore stages and isolated haustorial structures from infected wheat, generating 17,684 ESTs. We produced sequences from both the sexual (pycniospores, aeciospores and teliospores) and asexual (germinated urediniospores) stages of the life cycle. From pycniospores and aeciospores, produced by infecting the alternate host, meadow rue (Thalictrum speciosissimum), 4,869 and 1,292 reads were generated, respectively. We generated 3,703 ESTs from teliospores produced on the senescent primary wheat host. Finally, we generated 6,817 reads from haustoria isolated from infected wheat as well as 1,003 sequences from germinated urediniospores. Along with 25,558 previously generated ESTs, we compiled a database of 13,328 non-redundant sequences (4,506 singlets and 8,822 contigs). Fungal genes were predicted using the EST version of the self-training GeneMarkS algorithm. To refine the EST database, we compared EST sequences by BLASTN to a set of 454 pyrosequencing-generated contigs and Sanger BAC-end sequences derived both from the Pt genome, and to ESTs and genome reads from wheat. A collection of 6,308 fungal genes was identified and compared to sequences of the cereal rusts, Puccinia graminis f. sp. tritici (Pgt) and stripe rust, P. striiformis f. sp. tritici (Pst), and poplar leaf rust Melampsora species, and the corn smut fungus, Ustilago maydis (Um). While extensive homologies were found, many genes appeared novel and species-specific; over 40% of genes did not match any known sequence in existing databases. Focusing on spore stages, direct comparison to Um identified potential functional homologs, possibly allowing heterologous functional analysis in that model fungus. Many potentially secreted protein genes were identified by similarity searches against genes and proteins of Pgt and Melampsora spp., revealing apparent orthologs.
  
  
  CONCLUSIONS
  The current set of Pt unigenes contributes to gene discovery in this major cereal pathogen and will be invaluable for gene model verification in the genome sequence.
authors: Pacific Agri-Food Research Centre, Agriculture & Agri-Food Canada, Summerland, BC V0H 1Z0, Canada.
authors: Junhuan
authors: J
authors: Xu
authors: Rob
authors: R
authors: Linning
authors: John
authors: J
authors: Fellers
authors: Matthew
authors: M
authors: Dickinson
authors: Wenhan
authors: W
authors: Zhu
authors: Ivan
authors: I
authors: Antonov
authors: David L
authors: DL
authors: Joly
authors: Michael E
authors: ME
authors: Donaldson
authors: Tamar
authors: T
authors: Eilam
authors: Yehoshua
authors: Y
authors: Anikster
authors: Travis
authors: T
authors: Banks
authors: Sarah
authors: S
authors: Munro
authors: Michael
authors: M
authors: Mayo
authors: Brian
authors: B
authors: Wynhoven
authors: Johar
authors: J
authors: Ali
authors: Richard
authors: R
authors: Moore
authors: Brent
authors: B
authors: McCallum
authors: Mark
authors: M
authors: Borodovsky
authors: Barry
authors: B
authors: Saville
authors: Guus
authors: G
authors: Bakkeren
mesh_terms: D000465:Algorithms; D001487:Basidiomycota / Q000235:genetics* / Q000254:growth & development; D055028:Comparative Genomic Hybridization; D019295:Computational Biology; D030541:Databases, Genetic; D020224:Expressed Sequence Tags; D015723:Gene Library; D005800:Genes, Fungal; D023281:Genomics / Q000379:methods; D008969:Molecular Sequence Data; D012331:RNA, Fungal / Q000235:genetics; D017422:Sequence Analysis, DNA; D013172:Spores, Fungal / Q000235:genetics; D014908:Triticum / Q000382:microbiology; D003313:Zea mays / Q000382:microbiology
chemical_list: D012331:RNA, Fungal