Social Impact StatementAlthough tremendously successful at feeding humanity, row crop agriculture based on annuals contributes to numerous ecosystem dis‐services, ranging from soil degradation and aquatic eutrophication to greenhouse gas production. In contrast, perennial grain crops (which produce harvests for multiple seasons from single plantings) have the potential to provide valuable regulating and supporting ecosystem services in addition to food production. In particular, losses of ecological capital that threaten permanent food insecurity such as ~1% of global soil per year are expected to be mitigated or even reversed by crops that combine the high yield realized by scientific breeding via multiple cropping cycles from single plantings.Summary Perennial herbaceous may provide food and biomass while preserving ecological capital and reducing energy inputs. Sorghum has two perennial relatives and rich morphological diversity being used to breed for perenniality. We elucidate genetic determinants of rhizomatousness and survival, in two BC1F2 populations totaling 246 genotypes derived from backcrossing different annual Sorghum bicolor X perennial S. halepense F1 plants to a tetraploidized S. bicolor. RNA‐seq assisted in identifying candidate genes for rhizomatousness. Correspondence of rhizomatousness quantitative trait loci (QTLs) with those from two populations derived from crosses between S. halepense progenitors S. bicolor X S. propinquum suggests either the preservation of interspecific polymorphism or the formation of novel alleles following polyploid S. halepense formation. Correspondence of tillering and branching QTLs further supports their developmental. Identification of genes from RNA‐seq study within QTL intervals provides insight toward discovery of causal rhizomatous genes. An unexpected finding from both S. halepense‐ and S. propinquum‐derived populations is that alleles contributing to late flowering are related to reduced rhizomatousness. Twelve of 16 QTL regions conferring rhizomatousness fall in paleo‐duplicated regions tracing to single ancestral regions 96 million years ago, indicating that corresponding genes in these regions have retained similar functions since the duplication event.

KEY MESSAGE:Comparing populations derived, respectively, from polyploid Sorghum halepense and its progenitors improved knowledge of plant architecture and showed that S. halepense harbors genetic novelty of potential value for sorghum improvement Vegetative growth and the timing of the vegetative-to-reproductive transition are critical to a plant's fitness, directly and indirectly determining when and how a plant lives, grows and reproduces. We describe quantitative trait analysis of plant height and flowering time in the naturally occurring tetraploid Sorghum halepense, using two novel BC1F2 populations totaling 246 genotypes derived from backcrossing two tetraploid Sorghum bicolor x S. halepense F1 plants to a tetraploidized S. bicolor. Phenotyping for two years each in Bogart, GA and Salina, KS allowed us to dissect variance into narrow-sense genetic (QTLs) and environmental components. In crosses with a common S. bicolor BTx623 parent, comparison of QTLs in S. halepense, its rhizomatous progenitor S. propinquum and S. bicolor race guinea which is highly divergent from BTx623 permit inferences of loci at which new alleles have been associated with improvement of elite sorghums. The relative abundance of QTLs unique to the S. halepense populations may reflect its polyploidy and subsequent 'diploidization' processes often associated with the formation of genetic novelty, a possibility further supported by a high level of QTL polymorphism within sibling lines derived from a common S. halepense parent. An intriguing hypothesis for further investigation is that polyploidy of S. halepense following 96 million years of abstinence, coupled with natural selection during its spread to diverse environments across six continents, may provide a rich collection of novel alleles that offer potential opportunities for sorghum improvement.

Tillering and secondary branching are two plastic traits with high agronomic importance, especially in terms of the ability of plants to adapt to changing environments. We describe a quantitative trait analysis of tillering and secondary branching in two novel BC1F2 populations totaling 246 genotypes derived from backcrossing two Sorghum bicolor x S. halepense F1 plants to a tetraploidized S. bicolor. A two-year, two-environment phenotypic evaluation in Bogart, GA and Salina, KS permitted us to identify major effect and environment specific QTLs. Significant correlation between tillering and secondary branching followed by discovery of overlapping sets of QTLs continue to support the developmental relationship between these two organs and suggest the possibility of pleiotropy. Comparisons with two other populations sharing S. bicolor BTx623 as a common parent but sampling the breadth of the Sorghum genus, increase confidence in QTL detected for these two plastic traits and provide insight into the evolution of morphological diversity in the Eusorghum clade. Correspondence between flowering time and vegetative branching supports other evidence in suggesting a pleiotropic effect of flowering genes. We propose a model to predict biomass weight from plant architecture related traits, quantifying contribution of each trait to biomass and providing guidance for future breeding experiments.

From noble beginnings as a prospective forage, polyploid Sorghum halepense (‘Johnsongrass’) is both an invasive species and one of the world’s worst agricultural weeds. Formed by S. bicolor x S. propinquum hybridization, we show S. halepense to have S. bicolor-enriched allele composition and striking mutations in 5,957 genes that differentiate it from representatives of its progenitor species and an outgroup. The spread of S. halepense may have been facilitated by introgression from closely-related cultivated sorghum near genetic loci affecting rhizome development, seed size, and levels of lutein, a photochemical protectant and abscisic acid precursor. Rhizomes, subterranean stems that store carbohydrates and spawn clonal propagules, have growth correlated with reproductive rather than other vegetative tissues, and increase survival of both temperate cold seasons and tropical dry seasons. Rhizomes of S. halepense are more extensive than those of its rhizomatous progenitor S. propinquum, with gene expression including many alleles from its non-rhizomatous S. bicolor progenitor. The first surviving polyploid in its lineage in ∼96 million years, its post-Columbian spread across six continents carried rich genetic diversity that in the United States has facilitated transition from agricultural to non-agricultural niches. Projected to spread another 200–600 km northward in the coming century, despite its drawbacks S. halepense may offer novel alleles and traits of value to improvement of sorghum.

Despite a "ploidy barrier," interspecific crosses to wild and/or cultivated sorghum (Sorghum bicolor, 2n = 2x = 20) may have aided the spread across six continents of Sorghum halepense, also exemplifying risks of "transgene escape" from crops that could make weeds more difficult to control. Genetic maps of two BC1F1 populations derived from crosses of S. bicolor (sorghum) and S. halepense with totals of 722 and 795 single nucleotide polymorphism (SNP) markers span 37 and 35 linkage groups, with 2-6 for each of the 10 basic sorghum chromosomes due to fragments covering different chromosomal portions or independent segregation from different S. halepense homologs. Segregation distortion favored S. halepense alleles on chromosomes 2 (1.06-4.68 Mb, near a fertility restoration gene), 7 (1.20-6.16 Mb), 8 (1.81-5.33 Mb, associated with gene conversion), and 9 (47.5-50.1 Mb); and S. bicolor alleles on chromosome 6 (0-40 Mb), which contains both a large heterochromatin block and the Ma1 gene. Regions of the S. halepense genome that are recalcitrant to gene flow from sorghum might be exploited as part a multi-component system to reduce the likelihood of spread of transgenes or other modified genes. Its SNP profile suggests that chromosome segments from its respective progenitors S. bicolor and Sorghum propinquum have extensively recombined in S. halepense. This study reveals genomic regions that might discourage crop-to-weed gene escape, and provides a foundation for marker-trait association analysis to determine the genetic control of traits contributing to weediness, invasiveness, and perenniality of S. halepense.

Biofuel made from agricultural products has the potential in contribute to a stable supply of fuel for growing energy demands. Some salient plant traits, such as stem diameter and water content, and their relationship to other important biomass-related traits are so far poorly understood. Here, we performed QTL mapping for three stem diameter and two water content traits in a S. bicolor BTx623 x IS3620c recombinant inbred line population of 399 genotypes, and validated the genomic regions identified using genome-wide association studies (GWAS) in a diversity panel of 354 accessions. The discovery of both co-localized and non-overlapping loci affecting stem diameter traits suggests that stem widths at different heights share some common genetic control, but also have some distinct genetic influences. Co-localizations of stem diameter and water content traits with other biomass traits including plant height, flowering time and the ‘dry’ trait, suggest that their inheritance may be linked functionally (pleiotropy) or physically (linkage disequilibrium). Water content QTL in homeologous regions resulting from an ancient duplication event may have been retained and continue to have related functions for an estimated 96 million years. Integration of QTL and GWAS data advanced knowledge of the genetic basis of stem diameter and water content components in sorghum, which may lead to tools and strategies for either enhancing or suppressing these traits, supporting advances toward improved quality of plant-based biomass for biofuel production.

We describe a genetic map with a total of 381 bins of 616 genotyping by sequencing (GBS)-based SNP markers in a F6-F8 recombinant inbred line (RIL) population of 393 individuals derived from crossing S. bicolor BTx623 to S. bicolor IS3620C, a guinea line substantially diverged from BTx623. Five segregation distorted regions were found with four showing enrichment for S. bicolor alleles, suggesting possible selection during formation of this RIL population. A quantitative trait locus (QTL) study with this number of individuals, tripled relative to prior studies of this cross, provided resources, validated previous findings, and demonstrated improved power to detect plant height and flowering time related QTL relative to other published studies. An unexpected low correlation between flowering time and plant height permitted us to separate QTL for each trait and provide evidence against pleiotropy. Ten non- random syntenic regions conferring QTL for the same trait suggest that those QTL may represent alleles at genes functioning in the same manner since the 96 million year ago genome duplication that created these syntenic relationships, while syntenic regions conferring QTL for different trait may suggest sub-functionalization after duplication. Collectively, this study provides resources for marker-assisted breeding, as well as a framework for fine mapping and subsequent cloning of major genes for important traits such as plant height and flowering time in sorghum.

Primitive and exotic accessions of cotton are potential sources of favorable alleles for genetic improvement, enriching diversity in the genetically constricted gene pool of elite cultivars. Three exotic accessions of cotton (MDN101, MDN063 and MDN257), collected from different parts of Central America and converted to day-neutral flowering; and four elite cultivars (PD94042, DES56, PMHS200 and Acala Maxxa) representing the US cotton gene pool were used as parents to create experimental populations. The corresponding F2 and F2:3 progenies of these populations were grown in two successive years (i.e., some in 2011–2012, some in 2012–2013) and phenotypes were scored in both F2 and F2:3 progenies in all 3 years (2011–2012–2013). These populations were screened with 113 polymorphic microsatellite markers selected from “hotspots” for fiber quality quantitative trait loci in the cotton genome and single marker analyses were performed to identify significant associations of the markers with six fiber quality traits. A total of 134 nominal marker-trait associations were identified, among which 15 were significant after Bonferroni correction for multiple comparisons. In 67 of 134 nominal associations and 4 of 15 significant associations, the exotic parents contributed favorable alleles to multiple backgrounds and for multiple traits, in addition to the traits for which they were selected. These results indicate that utilization of exotic and wild accessions of cotton is useful in introducing favorable alleles into the cultivated cotton gene pool for genetic improvement.

Johnsongrass (Sorghum halepense) is a striking example of a post-Columbian founder event. This natural experiment within ecological time-scales provides a unique opportunity for understanding patterns of continent-wide genetic diversity following range expansion. Microsatellite markers were used for population genetic analyses including leaf-optimized Neighbor-Joining tree, pairwise FST, mismatch analysis, principle coordinate analysis, Tajima's D, Fu's F and Bayesian clusterings of population structure. Evidence indicates two geographically distant introductions of divergent genotypes, which spread across much of the US in <200 years. Based on geophylogeny, gene flow patterns can be inferred to have involved five phases. Centers of genetic diversity have shifted from two introduction sites separated by ~2000 miles toward the middle of the range, consistent with admixture between genotypes from the respective introductions. Genotyping provides evidence for a 'habitat switch' from agricultural to non-agricultural systems and may contribute to both Johnsongrass ubiquity and aggressiveness. Despite lower and more structured diversity at the invasion front, Johnsongrass continues to advance northward into cooler and drier habitats. Association genetic approaches may permit identification of alleles contributing to the habitat switch or other traits important to weed/invasive management and/or crop improvement.

PREMISE OF THE STUDY:Rhizomes, subterranean stems that grow horizontally, are a storage organ that is highly associated with overwintering and regrowth. This quantitative study aimed to discover genetic determinants of rhizomatousness, an important trait related to perenniality and invasiveness.METHODS:A population of 161 individuals of a recombinant inbred line (RIL) derived from morphologically distinct parents, Sorghum bicolor and Sorghum propinquum, which segregates for rhizomatousness, was phenotyped and genetically mapped.KEY RESULTS:Seven genomic regions influenced rhizomatousness in this population; four were "consensus" regions that correspond with previously detected quantitative trait loci (QTLs) in an F2 population of the same pedigree and with different levels of vegetative branching. Because rhizomatousness is a plastic trait that is greatly influenced by environment, overlap between regions discovered in the RIL and F2 populations validates the position and effect of QTLs. Correspondence with regions influencing vegetative branching indicates that some genes and biochemical pathways may influence both vegetative branching and rhizomatousness, while genes influencing only one trait may confer divergent aspects of development of these organs.CONCLUSIONS:Identifying genes conferring rhizomatousness and understanding their functions may provide opportunities to regulate plant growth for diverse applications. Increasing rhizomatousness may promote the productivity and perenniality of many grasses, especially biomass-dedicated crops, while decreasing rhizomatousness may improve monocarpic grain production and offer means to control many noxious weeds.

The complex genetic and environmental control of lint fiber yield and quality of cotton has long motivated interest in whether information from genetically-simpler trichome variations might contribute knowledge salient to cotton improvement. To investigate this question, from 3164 M5 lines resulting from EMS mutagenesis of two Gossypium hirsutum breeding lines, TAM 94L25 and Acala 1517-99, 106 lines with leaf and stem trichome variations and 55 control lines were further studied to investigate associations between trichome variation and lint fiber development. Although only weak correlation was found between stem/leaf trichome and fiber traits, we still found that among nine fiber traits measured in replicated trials, lines with mutations affecting stem trichome development had significant alterations for seven traits in the TAM 94L25 mutants, and six in the Acala 1517-99 mutants. While the small number of leaf trichome mutants found offered only minimal statistical power to resolve differences, mutant lines had significant alterations for three of the nine traits in the TAM 94L25 mutants, and two in the Acala 1517-99 mutants. In summary, mutants in leaf and/or stem trichome development often have altered lint fiber characteristics, supporting the hypothesis that there is considerable overlap in the sets of genetic factors acting in the development of these analogous organs. Moreover, visual selection of trichome mutants may be an effective screen to identify potential new alleles affecting lint fiber development.

An introduced species must contend with enormous environmental variation in its introduced range. In this study, we use niche models and ordination analyses to reconstruct changes in genotype, phenotype, and climatic niche of Johnsongrass Sorghum halepense, which is regarded as one of the world's most threatening invasive plants. In the United States, Johnsongrass has rapidly evolved within- and among-population genetic diversity; our results show that genetic differentiation in expanding Johnsongrass populations has resulted in phenotypic variation that is consistent with habitat and climatic variation encountered during its expansion. Moreover, Johnsongrass expanded from agricultural to non-agricultural habitat, and now, despite occupying overlapping ranges, extant agricultural and non-agricultural populations are genetically and phenotypically distinct and manifest different plastic responses when encountering environmental variation. Non-agricultural accessions are broadly distributed in climatic and geographic space and their fitness traits demonstrate plastic responses to common garden conditions that are consistent with local specialization. In contrast, agricultural accessions demonstrate general purpose' plastic responses and have more restricted climatic niches and geographic distributions. They also grow much larger than non-agricultural accessions. If these differences are adaptive, our results suggest that adaptation to local habitat variation plays a crucial role in the ecology of this invader. Further, its success relates to its ability to succeed on dual fronts, by responding simultaneously to habitat and climate variability and by capitalizing on differential responses to these factors during its range expansion.

Background: Domestication has played an important role in shaping characteristics of the inflorescence and plant height in cultivated cereals. Taking advantage of meta-analysis of QTLs, phylogenetic analyses in 502 diverse sorghum accessions, GWAS in a sorghum association panel (n = 354) and comparative data, we provide insight into the genetic basis of the domestication traits in sorghum and rice.Results: We performed genome-wide association studies (GWAS) on 6 traits related to inflorescence morphology and 6 traits related to plant height in sorghum, comparing the genomic regions implicated in these traits by GWAS and QTL mapping, respectively. In a search for signatures of selection, we identify genomic regions that may contribute to sorghum domestication regarding plant height, flowering time and pericarp color. Comparative studies across taxa show functionally conserved 'hotspots' in sorghum and rice for awn presence and pericarp color that do not appear to reflect corresponding single genes but may indicate co-regulated clusters of genes. We also reveal homoeologous regions retaining similar functions for plant height and flowering time since genome duplication an estimated 70 million years ago or more in a common ancestor of cereals. In most such homoeologous QTL pairs, only one QTL interval exhibits strong selection signals in modern sorghum.Conclusions: Intersections among QTL, GWAS and comparative data advance knowledge of genetic determinants of inflorescence and plant height components in sorghum, and add new dimensions to comparisons between sorghum and rice.

Seed size is closely related to fitness of wild plants, and its modification has been a key recurring element in domestication of seed/grain crops. In sorghum, a genomic and morphological model for panicoid cereals, a rich history of research into the genetics of seed size is reflected by a total of 13 likelihood intervals determined by conventional QTL (linkage) mapping in 11 nonoverlapping regions of the genome. To complement QTL data and investigate whether the discovery of seed size QTL is approaching "saturation," we compared QTL data to GWAS for seed mass, seed length, and seed width studied in 354 accessions from a sorghum association panel (SAP) that have been genotyped at 265,487 SNPs. We identified nine independent GWAS-based "hotspots" for seed size associations. Targeted resequencing near four association peaks with the most notable linkage disequilibrium provides further support of the role(s) of these regions in the genetic control of sorghum seed size and identifies two candidate causal variants with nonsynonymous mutations. Of nine GWAS hotspots in sorghum, seven have significant correspondence with rice QTL intervals and known genes for components of seed size on orthologous chromosomes. Identifying intersections between positional and association genetic data are a potentially powerful means to mitigate constraints associated with each approach, and nonrandom correspondence of sorghum (panicoid) GWAS signals to rice (oryzoid) QTL adds a new dimension to the ability to leverage genetic data about this important trait across divergent plants.

Genetic improvements for many fiber traits are obtained by mutagenesis of elite cottons, mitigating genetic uniformity in this inbred polyploid by contributing novel alleles important to ongoing crop improvement.

We identified quantitative trait loci influencing plant architecture that may be valuable in breeding of optimized genotypes for sustainable food and/or cellulosic biomass production, and advancing resilience to changing climates.

We describe a recombinant inbred line (RIL) population of 161 F5 genotypes for the widest euploid cross that can be made to cultivated sorghum (Sorghum bicolor) using conventional techniques, S. bicolor × Sorghum propinquum, that segregates for many traits related to plant architecture, growth and development, reproduction, and life history. The genetic map of the S. bicolor × S. propinquum RILs contains 141 loci on 10 linkage groups collectively spanning 773.1 cM. Although the genetic map has DNA marker density well-suited to quantitative trait loci mapping and samples most of the genome, our previous observations that sorghum pericentromeric heterochromatin is recalcitrant to recombination is highlighted by the finding that the vast majority of recombination in sorghum is concentrated in small regions of euchromatin that are distal to most chromosomes. The advancement of the RIL population in an environment to which the S. bicolor parent was well adapted (indeed bred for) but the S. propinquum parent was not largely eliminated an allele for short-day flowering that confounded many other traits, for example, permitting us to map new quantitative trait loci for flowering that previously eluded detection. Additional recombination that has accrued in the development of this RIL population also may have improved resolution of apices of heterozygote excess, accounting for their greater abundance in the F5 than the F2 generation. The S. bicolor × S. propinquum RIL population offers advantages over early-generation populations that will shed new light on genetic, environmental, and physiological/biochemical factors that regulate plant growth and development.

Suppression of seed shattering was a key step during crop domestication that we have previously suggested to be convergent among independent cereal lineages. Positional, association, expression, and mutant complementation data all implicate a WRKY transcription factor, SpWRKY, in conferring shattering to a wild sorghum relative, Sorghum propinquum. We hypothesize that SpWRKY functions in a manner analogous to Medicago and Arabidopsis homologs that regulate cell wall biosynthesis genes, with low expression toward the end of floral development derepressing downstream cell wall biosynthesis genes to allow deposition of lignin that initiates the abscission zone in the seed-pedicel junction. The recent discovery of a YABBY locus that confers shattering within Sorghum bicolor and other cereals validated our prior hypothesis that some parallel domestication may have been convergent. Ironically, however, the shattering allele of SpWRKY appears to be recently evolved in S. propinquum and illustrates a case in which the genetic control of a trait in a wild relative fails to extrapolate even to closely related crops. Remarkably, the SpWRKY and YABBY loci lie only 300 kb apart and may have appeared to be a single genetic locus in some sorghum populations.

For lignocellulosic bioenergy to be economically viable, genetic improvements must be made in feedstock quality including both biomass total yield and conversion efficiency. Toward this goal, multiple studies have considered candidate genes and discovered quantitative trait loci (QTL) associated with total biomass accumulation and/or grain production in bioenergy grass species including maize and sorghum. However, very little research has been focused on genes associated with increased biomass conversion efficiency. In this study, Trichoderma viride fungal cellulase hydrolysis activity was measured for lignocellulosic biomass (leaf and stem tissue) obtained from individuals in a F5 recombinant inbred Sorghum bicolor × Sorghum propinquum mapping population. A total of 49 QTLs (20 leaf, 29 stem) were associated with enzymatic conversion efficiency. Interestingly, six high-density QTL regions were identified in which four or more QTLs overlapped. In addition to enzymatic conversion efficiency QTLs, two QTLs were identified for biomass crystallinity index, a trait which has been shown to be inversely correlated with conversion efficiency in bioenergy grasses. The identification of these QTLs provides an important step toward identifying specific genes relevant to increasing conversion efficiency of bioenergy feedstocks. DNA markers linked to these QTLs could be useful in marker-assisted breeding programs aimed at increasing overall bioenergy yields concomitant with selection of high total biomass genotypes.

To explore types, levels and patterns of genetic divergence among diploid Gossypium (cotton) genomes, 780 cDNA, genomic DNA and simple sequence repeat (SSR) loci were re-sequenced in G o ssypium herbaceum (A1 genome), G. arboreum (A2), G. raimondii (D5), G. trilobum (D8), G. sturtianum (C1) and an outgroup, Gossypioides kirkii . Divergence among these genomes ranged from 7.32 polymorphic base pairs per 100 between G. kirkii and G. herbaceum (A1) to only 1.44 between G . herbaceum (A1) and G. arboreum (A2). SSR loci are least conserved with 12.71 polymorphic base pairs and 3.77 polymorphic sites per 100 base pairs, whereas expressed sequence tags are most conserved with 3.96 polymorphic base pairs and 2.06 sites. SSR loci also exhibit the highest percentage of ‘extended polymorphisms’ (spanning multiple consecutive nucleotides). The A genome lineage was particularly rapidly evolving, with the D genome also showing accelerated evolution relative to the C genome. Unexpected asymmetry in mutation rates was found, with much more transition than transversion mutation in the D genome after its divergence from a common ancestor shared with the A genome. This large quantity of orthologous DNA sequence strongly supports a phylogeny in which A–C divergence is more recent than A–D divergence, a subject that is of much importance in view of A–D polyploid formation being key to the evolution of the most productive and finest-quality cottons. Loci that are monomorphic within A or D genome types, but polymorphic between genome types, may be of practical importance for identifying locus-specific DNA markers in tetraploid cottons including leading cultivars.