Program Book - 27th Fungal Genetics Conference

CONCURRENT SESSION ABSTRACTSA draft genome of the ectomycorrhizal fungus Rhizopogon vesiculosus: Characterization of mating system and heterozygosity within the dikaryon. AlijaMujic, Joseph Spatafora. Botany and Plant Pathology, Oregon State University, Corvallis, OR.Species of Rhizopogon are EM symbionts of trees in family Pinaceae and produce basidiospores within hypogeous false truffles that are dispersed bymycophagous mammals. All known members of R. subgenus Villosuli form obligate EM relationships with Pseudotsuga spp. (Douglas Fir) and are the onlymembers of the genus known to possess this host association. R. vesiculosus, along with its cryptic sister species R. vinicolor, possess a sympatricdistribution where sampled within the range of their host tree, P. menziesii. While the sporocarp and EM morphology of these fungi may be highly similar;they possess striking life history differences with R. vesiculosus producing larger vegetative genets and displaying greater population structure at bothlocal and landscape scales. We have sequenced the genome of R. vesiculosus using dikaryotic tissue and a whole genome shotgun sequencing approach onthe Illumina HiSeq platform. De novo assembly of the genome was performed using VELVET 1.19 and gene predictions were made using AUGUSTUS withLaccaria bicolor as a training model. The draft genome assembled to a total length of 46 Mb in 6700 contigs with an N50 of 26,783, a maximum contig sizeof 446,818 bp, and 12,604 predicted genes. Here we characterize the mating system of R. vesiculosus, which possesses both an A-locus encoding aheterodimer transcription factor, as well a B-locus encoding transmembrane pheromone receptors and pheromone precursor genes. We presentcomparisons of the mating system of R. vinicolor and its similarities to other members of Boletales (e.g., Serpula) and differences with Agaricales (e.g.,Laccaria). Due to the dikaryotic nature of the genome sequence produced for R. vesiculosus, single nucleotide polymorphisms (SNPs) can be observed andused to characterize allelic variation. SNPs observed in protein coding regions of both MAT loci indicate that R. vesisculosus is likely heterothallic. We havealso characterized heterozygosity across the whole genome in order to identify hypervariable regions. This genome will allow for comparative analysis ofgene content, mating type system with other Basidiomycota and, ultimately, for population/species-level genomic studies within Rhizopogon.Metatranscriptomic analysis of ectomycorrhizal root clusters in Pinus taeda: new methodologies for assessing functional gene expression in situ. H.-L.Liao 1 , Y. Chen 2 , T. D. Bruns 3 , K. G. Peay 4 , J. W. Taylor 3 , S. Branco 3 , J. M. Talbot 4 , R. Vilgalys 1 . 1) Department of Biology Duke University, Durham, NC; 2)School of Medicine, Duke University, Durham, NC; 3) Department of Plant and Microbial Biology, UC-Berkeley, Berkeley, CA; 4) Department of Biology,Stanford University, Stanford, CA.A highly diverse community of ectomycorrhizal (ECM) fungi are known to associate with members of the genus Pinus. Less is known about how diversefungal communities affect functional diversity within ECM roots. Here we present an optimized method for metatranscriptomic analysis of the ECM-pineroot interaction in a natural system. RNA was purified using a CTAB method from individual ECM root clusters collected at varying spatial scales across thedistribution range of P. taeda, and sequenced using Illumina HiSeq technology. About 35 million qualified reads were obtained. Sequences were initiallyassembled using reference based mapping (Bowtie) to sort the reads that represent rRNA from fungal and bacterial species. Reads from divergent regions(D1-D2) of fungal LSU rRNA were used to identify dominant ECM and other fungal community members. Subsequently, P. taeda genes and functionalgenes of dominant fungal species were sorted using public cDNA databases. The Trinity package was used for de novo assembly of un-mapped reads(mostly fungal genes). Blastx and Go packages were used for gene annotation. A typical ECM root cluster was found 45% P. taeda genes, 3% fungal rRNA,0.05% bacterial 16S rRNA, 30% fungal functional genes, 10% unknown sequences, and 12% unassembled reads. Analysis of D1-D2 LSU sequencesconfirmed that a single ECM fungal species usually dominates individual root clusters. De novo assemblies of fungal genes yielded 120 thousand contigsfrom 10 million reads representing 90 thousand unique genes with highly similarity to known ECM fungi. Functional analysis revealed that most of thetranscripts recovered were involved with translation, protein degradation, heat shock, superoxide metabolism, electron transfer, signaling, and C/Nmetabolism. Highly expressed transcripts recovered from Piloderma, which was abundant in our samples, included genes encoding a wide array ofmetabolic enzymes: chitosanase, phosphatase, glutamine synthetase, terpene synthases, b-glucanase; transporters for P+ and oligopeptides; cell signaling:calmodulin, cAMP-regulated phosphoprotein (Igo1); C/N related genes: lectin, cross-pathway control (cpc1); as well as several genes with unknownfunction. Future studies will seek to address how ECM metatranscriptomes change in response to different Pinus hosts and across different spatial scales.27th Fungal Genetics Conference | 41