Next Generation Sequencing On Species Important to the Turfgrass Industry.

The bentgrasses (Agrostis spp.) are important species to the turfgrass industry because of their unique growth and aesthetic characteristics that make them ideally suited for use on golf course tees, fairways, and putting greens.  Molecular marker development is difficult in Agrostis due to the limited amount of available DNA sequence data.  For example there are 16,992 Agrostis EST sequences (as of April 23, 2009) published in the National Center for Biotechnology Information databases, dwarfed by the millions of EST sequences available for cereal grasses.  Illumina sequencing technology is among the most popular of the next generation sequencing technologies and provides an affordable way of producing large amounts of sequence data.  The objective of this study was to evaluate the Illumina Genome Analyzer for the generation of EST sequence data from one velvet bentgrass (A. canina L.), three creeping bentgrasses (A. stolonifera L.), two dollar spot causing fungal isolates (Sclerotinia homeocarpa FT Bennett), and one brown patch causing fungal isolate (Rhizoctonia solani Kuhn).   In addition, the feasibility of assembling the short Illumina sequencing reads into usable data was tested.  A total of 1,026,283 (Agrostis), 1,834,661 (Sclerotinia), and 326,174 (Rhizoctonia) 36-mer sequences were generated.  The software programs Edena and Velvet were used to assemble the sequences into contigs.  A total of 134 contigs were assembled for the turfgrass libraries, 857 for the Sclerotinia libraries, and 62 for the Rhizoctonia library.  While this sequencing run generated approximately 10 percent of the expected amount of data, more than 1000 EST sequences were recovered.  This preliminary experiment demonstrates the utility of high throughput sequencing on species important to the turfgrass industry and the need for additional sequencing.