De Novo Hybrid Transcriptome Assembly and RNA-Seq Analysis of Switchgrass Crowns and Rhizomes

Switchgrass is a perennial native grass species which has been selected for use as a lignocellulosic bioenergy feedstock.  Improving yield and conversion are well studied key traits for feedstock plants.  However, an equally important but poorly studied trait is the ability to regrow tillers in the spring without having significant winter kill.  The initiation point of this regrowth is the below-ground crown and rhizome tissues of the plant and investigation of these tissues should provide insight on factors involved perenniality and winter survivability.  An RNA-Seq approach was taken to analyze the gene expression differences in crown and rhizome tissues between two switchgrass populations with contrasting winter survivability.  De novo assembly was utilized to generate a reference transcriptome to use for gene expression analysis.  A hybrid assembly using transcriptome sequence acquired from 454 (1.3 million reads) as well as Illumina (100 million reads) platforms was generated using the software programs Velvet and Oases.  This assembly was utilized as a reference for RNA-Seq analysis using three biological replicates (~25 million 75 bp reads each) from each switchgrass cultivar harvested on the same date in September 2010.  Gene expression differences between the two cultivars at this specific harvest date were documented.  Ongoing RNA-Seq studies using more populations and samples harvested at specific developmental stages can be expected to provide greater insights on transcriptomic changes as related to winter survivability.