1 Department of Molecular Biology and Genetics - Afgrødegenetik og Bioteknologi, Department of Molecular Biology and Genetics, Science and Technology, Aarhus University2 Plant Sciences Unit, Institute for Agricultural and Fisheries Research (ILVO)3 Department of Molecular Biology and Genetics - Afgrødegenetik og Bioteknologi, Department of Molecular Biology and Genetics, Science and Technology, Aarhus University
To advance the application of molecular breeding in Lolium perenne, we have generated a sequence resource to facilitate gene discovery and SNP marker development. Illumina GAII transcriptome sequencing was performed on meristem-enriched samples of 14 Lolium genotypes. De novo assemblies for individual genotypes was carried out with 12-30M paired-end reads per genotype using the CLCBio Genomics Workbench, yielding around 70,000 contigs per genotype. 54% of these contigs have a significant BLAST hit with a predicted Brachypodium gene. Vice versa, for around 60% of the 26,552 predicted Brachypodium genes a homologous Lolium sequence has been identified. We are subsequently merging these 14 individual transcriptomes to a common reference transcriptome for Lolium. The data resource is currently functional for interrogation of individual candidate genes. Through phylogenetic analysis using predicted genes from seven completely sequenced genomes together with the Lolium contigs, orthologous groups are established and most probable candidate orthologues in Lolium are assigned. We are currently validating the de novo transcriptome assembly by Sanger sequencing and will develop and validate a set of SNP markers in selected candidate genes. In parallel, a germplasm collection of 602 Lolium genotypes was established and is being phenotyped for plant architecture, reproductive characteristics, flowering time, and forage quality traits. We will test through association genetics whether phenotypic variation can be explained by SNP polymorphisms in selected candidate genes. Information on significant associations between SNPs and investigated traits will eventually be used to design appropriate crossing and selection schemes using selected subsets of genotypes from this collection.