Identification and Analysis of Cell Wall-related Genes in Switchgrass

Switchgrass (Panicum virgatum), a C4 perennial grass, is of research interest due to its potential as a dedicated bioenergy crop. Whole genome sequencing of P. virgatum L., clone AP13, is underway at the Joint Genome Institute in collaboration with the Joint BioEnergy Institute and other Bioenergy Research Centers. Switchgrass AP10 is tetraploid and carries four copies of most alleles. To facilitate assembly of the AP10 switchgrass genome, we constructed two Bacterial Artificial Chromosome libraries. We are using these libraries and comparative genomics approaches to identify full-length sequences of the genes critical for cell wall biosynthesis/modification and stress-associated responses in switchgrass.

In collaboration with JGI, we have fully sequenced 310 BAC clones containing more than 1000 genes with predicted roles in cell wall biogenesis/degradation, assembly/remodeling and stress associated responses. These include glycosyl transfereases, glycosyl hydrolases, kinases, transcription factors, transporters etc. Q-PCR-based gene expression analysis of representative cellulose synthase/cellulose synthase-like genes among diverse switchgrass tissues revealed conserved as well as diversified gene expression patterns.

The tetraploid nature of switchgrass AP10 and the polymorphism of each subgenome makes it difficult to distinguish gene clusters corresponding to each subgenome. For this reason, researchers currently have little information about what distinguishes each subgenome, complicating assembly. To address this issue, we analyzed the switchgrass Cellulose synthase-like F (CslF) region. CslF genes are involved in synthesis of mixed-linkage glucans in grasses. In rice, six CslF genes are present in a cluster on chromosome 7. We identified the corresponding syntenic region in switchgrass and found that the six CslF genes are present on a 191 Kb genomic fragment and that the gene order is conserved between rice and switchgrass. We identified 15 BAC clones carrying the CslF gene cluster.  Some of the BAC end sequences were overlapping but were not identical. We then fully sequenced these BAC clones. Assembly of these clones resulted in two contigs.