Five nuclear loci resolve the polyploid history of switchgrass (Panicum virgatum L.) and relatives.
ABSTRACT: Polyploidy poses challenges for phylogenetic reconstruction because of the need to identify and distinguish between homoeologous loci. This can be addressed by use of low copy nuclear markers. Panicum s.s. is a genus of about 100 species in the grass tribe Paniceae, subfamily Panicoideae, and is divided into five sections. Many of the species are known to be polyploids. The most well-known of the Panicum polyploids are switchgrass (Panicum virgatum) and common or Proso millet (P. miliaceum). Switchgrass is in section Virgata, along with P. tricholaenoides, P. amarum, and P. amarulum, whereas P. miliaceum is in sect. Panicum. We have generated sequence data from five low copy nuclear loci and two chloroplast loci and have clarified the origin of P. virgatum. We find that all members of sects. Virgata and Urvilleana are the result of diversification after a single allopolyploidy event. The closest diploid relatives of switchgrass are in sect. Rudgeana, native to Central and South America. Within sections Virgata and Urvilleana, P. tricholaenoides is sister to the remaining species. Panicum racemosum and P. urvilleanum form a clade, which may be sister to P. chloroleucum. Panicum amarum, P. amarulum, and the lowland and upland ecotypes of P. virgatum together form a clade, within which relationships are complex. Hexaploid and octoploid plants are likely allopolyploids, with P. amarum and P. amarulum sharing genomes with P. virgatum. Octoploid P. virgatum plants are formed via hybridization between disparate tetraploids. We show that polyploidy precedes diversification in a complex set of polyploids; our data thus suggest that polyploidy could provide the raw material for diversification. In addition, we show two rounds of allopolyploidization in the ancestry of switchgrass, and identify additional species that may be part of its broader gene pool. This may be relevant for development of the crop for biofuels.
Project description:Geographic patterns of genetic variation are shaped by multiple evolutionary processes, including genetic drift, migration and natural selection. Switchgrass (Panicum virgatum L.) has strong genetic and adaptive differentiation despite life history characteristics that promote high levels of gene flow and can homogenize intraspecific differences, such as wind-pollination and self-incompatibility. To better understand how historical and contemporary factors shape variation in switchgrass, we use genotyping-by-sequencing to characterize switchgrass from across its range at 98 042 SNPs. Population structuring reflects biogeographic and ploidy differences within and between switchgrass ecotypes and indicates that biogeographic history, ploidy incompatibilities and differential adaptation each have important roles in shaping ecotypic differentiation in switchgrass. At one extreme, we determine that two Panicum taxa are not separate species but are actually conspecific, ecologically divergent types of switchgrass adapted to the extreme conditions of coastal sand dune habitats. Conversely, we identify natural hybrids among lowland and upland ecotypes and visualize their genome-wide patterns of admixture. Furthermore, we determine that genetic differentiation between primarily tetraploid and octoploid lineages is not caused solely by ploidy differences. Rather, genetic diversity in primarily octoploid lineages is consistent with a history of admixture. This suggests that polyploidy in switchgrass is promoted by admixture of diverged lineages, which may be important for maintaining genetic differentiation between switchgrass ecotypes where they are sympatric. These results provide new insights into the mechanisms shaping variation in widespread species and provide a foundation for dissecting the genetic basis of adaptation in switchgrass.
Project description:BACKGROUND: Karyotypes can provide information about taxonomic relationships, genetic aberrations, and the evolutionary origins of species. However, differentiation of the tiny chromosomes of switchgrass (Panicum virgatum L.) and creation of a standard karyotype for this bioenergy crop has not been accomplished due to lack of distinguishing features and polyploidy. RESULTS: A cytogenetic study was conducted on a dihaploid individual (2n?=?2X?=?18) of switchgrass to establish a chromosome karyotype. Size differences, condensation patterns, and arm-length ratios were used as identifying features and fluorescence in-situ hybridization (FISH) assigned 5S and 45S rDNA loci to chromosomes 7 and 2 respectively. Both a maize CentC and a native switchgrass centromeric repeat (PviCentC) that shared 73% sequence identity demonstrated a strong signal on chromosome 3. However, only the PviCentC probe labeled the centromeres of all chromosomes. Unexpected PviCentC and 5S rDNA hybidization patterns were consistent with severe reduction or total deletion of these repeats in one subgenome. These patterns were maintained in tetraploid and octoploid individuals. The 45S rDNA repeat produced the expected number of loci in dihaploid, tetraploid and octoploid individuals. Differences observed at the 5S rDNA loci between the upland and lowland ecotypes of switchgrass provided a basis for distinguishing these subpopulations. CONCLUSION: Collectively, these results provide a quantitative karyotype of switchgrass chromosomes. FISH analyses indicate genetic divergence between subgenomes and allow for the classification of switchgrass plants belonging to divergent genetic pools. Furthermore, the karyotype structure and cytogenetic analysis of switchgrass provides a framework for future genetic and genomic studies.
Project description:Switchgrass (Panicum virgatum L.) is a perennial grass that has been designated as an herbaceous model biofuel crop for the United States of America. To facilitate accelerated breeding programs of switchgrass, we developed both an association panel and linkage populations for genome-wide association study (GWAS) and genomic selection (GS). All of the 840 individuals were then genotyped using genotyping by sequencing (GBS), generating 350 GB of sequence in total. As a highly heterozygous polyploid (tetraploid and octoploid) species lacking a reference genome, switchgrass is highly intractable with earlier methodologies of single nucleotide polymorphism (SNP) discovery. To access the genetic diversity of species like switchgrass, we developed a SNP discovery pipeline based on a network approach called the Universal Network-Enabled Analysis Kit (UNEAK). Complexities that hinder single nucleotide polymorphism discovery, such as repeats, paralogs, and sequencing errors, are easily resolved with UNEAK. Here, 1.2 million putative SNPs were discovered in a diverse collection of primarily upland, northern-adapted switchgrass populations. Further analysis of this data set revealed the fundamentally diploid nature of tetraploid switchgrass. Taking advantage of the high conservation of genome structure between switchgrass and foxtail millet (Setaria italica (L.) P. Beauv.), two parent-specific, synteny-based, ultra high-density linkage maps containing a total of 88,217 SNPs were constructed. Also, our results showed clear patterns of isolation-by-distance and isolation-by-ploidy in natural populations of switchgrass. Phylogenetic analysis supported a general south-to-north migration path of switchgrass. In addition, this analysis suggested that upland tetraploid arose from upland octoploid. All together, this study provides unparalleled insights into the diversity, genomic complexity, population structure, phylogeny, phylogeography, ploidy, and evolutionary dynamics of switchgrass.
Project description:Panicum represents a large genus of many North American prairie grass species. These include switchgrass (Panicum virgatum), a biofuel crop candidate with wide geographic range, as well as Panicum hallii, a close relative to switchgrass, which serves as a model system for the study of Panicum genetics due to its diploid genome and short growth cycles. For the advancement of switchgrass as a biofuel crop, it is essential to understand host microbiome interactions, which can be impacted by plant genetics and environmental factors inducing ecotype-specific phenotypic traits. We here compared rhizosphere and root endosphere bacterial communities of upland and lowland P. virgatum and P. hallii genotypes planted at two sites in Texas. Our analysis shows that sampling site predominantly contributed to bacterial community variance in the rhizosphere, however, impacted root endosphere bacterial communities much less. Instead we observed a relatively large core endophytic microbiome dominated by ubiquitously root-colonizing bacterial genera Streptomyces, Pseudomonas, and Bradyrhizobium. Endosphere communities displayed comparable diversity and conserved community structures across genotypes of both Panicum species. Functional insights into interactions between P. hallii and its root endophyte microbiome could hence inform testable hypotheses that are relevant for the improvement of switchgrass as a biofuel crop.
Project description:Switchgrass (Panicum virgatum) is a polyploid, outcrossing grass species native to North America and has recently been recognized as a potential biofuel feedstock crop. Significant phenotypic variation including ploidy is present across the two primary ecotypes of switchgrass, referred to as upland and lowland switchgrass. The tetraploid switchgrass genome is approximately 1400 Mbp, split between two subgenomes, with significant repetitive sequence content limiting the efficiency of re-sequencing approaches for determining genome diversity. To characterize genetic diversity in upland and lowland switchgrass as a first step in linking genotype to phenotype, we designed an exome capture probe set based on transcript assemblies that represent approximately 50 Mb of annotated switchgrass exome sequences. We then evaluated and optimized the probe set using solid phase comparative genome hybridization and liquid phase exome capture followed by next-generation sequencing. Using the optimized probe set, we assessed variation in the exomes of eight switchgrass genotypes representing tetraploid lowland and octoploid upland cultivars to benchmark our exome capture probe set design. We identified ample variation in the switchgrass genome including 1,395,501 single nucleotide polymorphisms (SNPs), 8173 putative copy number variants and 3336 presence/absence variants. While the majority of the SNPs (84%) detected was bi-allelic, a substantial number was tri-allelic with limited occurrence of tetra-allelic polymorphisms consistent with the heterozygous and polyploid nature of the switchgrass genome. Collectively, these data demonstrate the efficacy of exome capture for discovery of genome variation in a polyploid species with a large, repetitive and heterozygous genome.
Project description:Although yield trials for switchgrass (Panicum virgatum L.), a potentially high value biofuel feedstock crop, are currently underway throughout North America, the genetic tools for crop improvement in this species are still in the early stages of development. Identification of high-density molecular markers, such as single nucleotide polymorphisms (SNPs), that are amenable to high-throughput genotyping approaches, is the first step in a quantitative genetics study of this model biofuel crop species. We generated and sequenced expressed sequence tag (EST) libraries from thirteen diverse switchgrass cultivars representing both upland and lowland ecotypes, as well as tetraploid and octoploid genomes. We followed this with reduced genomic library preparation and massively parallel sequencing of the same samples using the Illumina Genome Analyzer technology platform. EST libraries were used to generate unigene clusters and establish a gene-space reference sequence, thus providing a framework for assembly of the short sequence reads. SNPs were identified utilizing these scaffolds. We used a custom software program for alignment and SNP detection and identified over 149,000 SNPs across the 13 short-read sequencing libraries (SRSLs). Approximately 25,000 additional SNPs were identified from the entire EST collection available for the species. This sequencing effort generated data that are suitable for marker development and for estimation of population genetic parameters, such as nucleotide diversity and linkage disequilibrium. Based on these data, we assessed the feasibility of genome wide association mapping and genomic selection applications in switchgrass. Overall, the SNP markers discovered in this study will help facilitate quantitative genetics experiments and greatly enhance breeding efforts that target improvement of key biofuel traits and development of new switchgrass cultivars.
Project description:We report here the improved draft genome sequence of Bacillus sp. strain YF23, a bacterium originally isolated from switchgrass (Panicum virgatum) plants and shown to exhibit plant growth-promoting activity. The genome comprised 5.82 Mbp, containing 5,933 genes, with 193 as RNA genes, and a GC content of 35.10%.
Project description:We report here the genome assembly and analysis of Microbacterium strain sp. LKL04, a Gram-positive bacterial endophyte isolated from switchgrass plants (Panicum virgatum) grown on a reclaimed coal-mining site. The 2.9-Mbp genome of this bacterium was assembled into a single contig encoding 2,806 protein coding genes.
Project description:Background:Newly formed polyploids may experience short-term adaptative changes in their genome that may enhance the resistance of plants to stress. Considering the increasingly serious effects of drought on biofuel plants, whole genome duplication (WGD) may be an efficient way to proceed with drought resistant breeding. However, the molecular mechanism of drought response before/after WGD remains largely unclear. Result:We found that autoploid switchgrass (Panicum virgatum L.) 8X Alamo had higher drought tolerance than its parent amphidiploid 4X Alamo using physiological tests. RNA and microRNA sequencing at different time points during drought were then conducted on 8X Alamo and 4X Alamo switchgrass. The specific differentially expressed transcripts (DETs) that related to drought stress (DS) in 8X Alamo were enriched in ribonucleoside and ribonucleotide binding, while the drought-related DETs in 4X Alamo were enriched in structural molecule activity. Ploidy-related DETs were primarily associated with signal transduction mechanisms. Weighted gene co-expression network analysis (WGCNA) detected three significant DS-related modules, and their DETs were primarily enriched in biosynthesis process and photosynthesis. A total of 26 differentially expressed microRNAs (DEmiRs) were detected, and among them, sbi-microRNA 399b was only expressed in 8X Alamo. The targets of microRNAs that were responded to polyploidization and drought stress all contained cytochrome P450 and superoxide dismutase genes. Conclusions:This study explored the drought response of 8X and 4X Alamo switchgrass on both physiological and transcriptional levels, and provided experimental and sequencing data basis for a short-term adaptability study and drought-resistant biofuel plant breeding.
Project description:Hsp20 proteins exist in all plant species and represent the most abundant small heat shock proteins (sHSPs) in plants. Hsp20s were known as chaperones maintaining cellular homeostasis during heat or other kinds of abiotic stresses. The objective of this study was to understand the phylogenetic relationship, genomic organization, diversification of motif modules, genome localization, expression profiles, and interaction networks of switchgrass (Panicum virgatum L.) Hsp20s (PvHsp20s). A total of 63 PvHsp20s were identified with their consensus as well as unique ACD motifs and gene structures analyzed. Most PvHsp20s (87%) were responsive to heat and other kinds of abiotic stresses. When under optimum growth condition, 38 of them displayed relative higher expression levels in inflorescence and seeds, suggesting their protective roles in the stress-sensitive reproductive organs. An in silico analysis of interaction network of PvHsp20 proteins further revealed potential interactive proteins, including stress-inducible ones in the network. Furthermore, PvHsp20 genes unevenly distributed in two sets of homeologous chromosomes, and only segmental duplication was found among the paralogous gene pairs, reflecting that the allotetraploidization of switchgrass allowed the accumulation of PvHsp20s that in turn facilitated its successful adaptation in hot and dry plateaus of North America. The present results provided an insight into PvHsp20s with an emphasis on the uniqueness of this gene family in switchgrass. Such information shall also be useful in functional studies of PvHsp20 genes and molecular breeding of switchgrass.