Location: Grain, Forage & Bioenergy Research
2012 Annual Report
University of Nebraska scientists were primarily responsible for labeling plants and their analyses and sequencing. Plant materials were labeled with 13CO2 and 15N (urea) in spring FY11 were collected throughout the growing season, separated into respective plant parts, oven dried and ground. Several samples were analyzed by mass spectrometry to gauge the level of incorporation of 13C and 15N into plant parts. Initial analysis showed that plants were labeled adequately and there was an expected dilution of label during the period of active growth. Crown + rhizomes were harvested from 2 populations at selected times over the growing season, cleaned and flash frozen in liquid nitrogen. Roots were also collected from each sample at each harvest date. All frozen plant materials were stored at -80C for future RNA analyses by high-throughput sequencing. High-throughput sequencing of previously collected crown materials from cv Summer was performed using the Illumina Hi-Seq 2000 instrument. The ~ 1.2 billion sequences obtained from cv Summer crowns + rhizomes using are being analyzed.
In an experiment completed in late FY11, about ~200 million short read sequences were obtained through Illumina sequencing of triplicate samples of crowns + rhizomes obtained from cv Kanlow and cv Summer for a single harvest date in late September. These reads were mapped to the switchgrass draft genome released by the JGI in January 2012. These analyses have been completed and results indicate substantial and significant differences in the metabolism of the crowns and rhizomes obtained from cv Summer plants as compared to tissues obtained from cv Kanlow plants.
For Objective 2, Crowns and rhizomes were collected from several plants from an octaploid nursery containing plants divergently selected for forage digestibility and exhibiting differences in winter hardiness. Crowns and rhizomes will be collected from 44 plants post-frost fro transcriptomic analyses using the Illumina Hi-Seq system. These studies will augment ongoing genotype-by-sequencing work on the same plants.
For Objective 3, phenotyping of plants is proceeding and will be expected to be completed in the autumn of 2012. These plants have been processed for genotype-by sequencing.
Communication was by email and telephone between the ADODR and appropriate project personnel at distant locations (ARS-Albany and University of Nebraska-Kearney) and by in-person meetings for project personnel located in Lincoln, NE. All team members were cognizant of the planned experiments and had approved implementation.