Patricia E. Klein

The Sorghum bicolor genome and the diversification of grasses

Sorghum, an African grass related to sugar cane and maize, is grown for food, feed, fibre and fuel. We present an initial analysis of the ∼730-megabase Sorghum bicolor (L.) Moench genome, placing ∼98% of genes in their chromosomal context using whole-genome shotgun sequence validated by genetic, physical and syntenic information. Genetic recombination is largely confined to about one-third of the sorghum genome with gene order and density similar to those of rice. Retrotransposon accumulation in recombinationally recalcitrant heterochromatin explains the ∼75% larger genome size of sorghum compared with rice. Although gene and repetitive DNA distributions have been preserved since palaeopolyploidization ∼70 million years ago, most duplicated gene sets lost one member before the sorghum–rice divergence. Concerted evolution makes one duplicated chromosomal segment appear to be only a few million years old. About 24% of genes are grass-specific and 7% are sorghum-specific. Recent gene and microRNA duplications may contribute to sorghum’s drought tolerance.

Transcriptional Profiling of Sorghum Induced by Methyl Jasmonate, Salicylic Acid, and Aminocyclopropane Carboxylic Acid Reveals Cooperative Regulation and Novel Gene Responses

We have conducted a large-scale study of gene expression in the C4 monocot sorghum (Sorghum bicolor) L. Moench cv BTx623 in response to the signaling compounds salicylic acid (SA), methyl jasmonate (MeJA), and the ethylene precursor aminocyclopropane carboxylic acid. Expression profiles were generated from seedling root and shoot tissue at 3 and 27 h, using a microarray containing 12,982 nonredundant elements. Data from 102 slides and quantitative reverse transcription-PCR data on mRNA abundance from 171 genes were collected and analyzed and are here made publicly available. Numerous gene clusters were identified in which expression was correlated with particular signaling compound and tissue combinations. Many genes previously implicated in defense responded to the treatments, including numerous pathogenesis-related genes and most members of the phenylpropanoid pathway, and several other genes that may represent novel activities or pathways. Genes of the octadecanoic acid pathway of jasmonic acid (JA) synthesis were induced by SA as well as by MeJA. The resulting hypothesis that increased SA could lead to increased endogenous JA production was confirmed by measurement of JA content. Comparison of responses to SA, MeJA, and combined SA+MeJA revealed patterns of one-way and mutual antagonisms, as well as synergistic effects on regulation of some genes. These experiments thus help further define the transcriptional results of cross talk between the SA and JA pathways and suggest that a subset of genes coregulated by SA and JA may comprise a uniquely evolved sector of plant signaling responsive cascades.

Coincident light and clock regulation of pseudoresponse regulator protein 37 (PRR37) controls photoperiodic flowering in sorghum.

Optimal flowering time is critical to the success of modern agriculture. Sorghum is a short-day tropical species that exhibits substantial photoperiod sensitivity and delayed flowering in long days. Genotypes with reduced photoperiod sensitivity enabled sorghums utilization as a grain crop in temperate zones worldwide. In the present study, Ma1, the major repressor of sorghum flowering in long days, was identified as the pseudoresponse regulator protein 37 (PRR37) through positional cloning and analysis of SbPRR37 alleles that modulate flowering time in grain and energy sorghum. Several allelic variants of SbPRR37 were identified in early flowering grain sorghum germplasm that contain unique loss-of-function mutations. We show that in long days SbPRR37 activates expression of the floral inhibitor CONSTANS and represses expression of the floral activators Early Heading Date 1, FLOWERING LOCUS T, Zea mays CENTRORADIALIS 8, and floral induction. Expression of SbPRR37 is light dependent and regulated by the circadian clock, with peaks of RNA abundance in the morning and evening in long days. In short days, the evening-phase expression of SbPRR37 does not occur due to darkness, allowing sorghum to flower in this photoperiod. This study provides insight into an external coincidence mechanism of photoperiodic regulation of flowering time mediated by PRR37 in the short-day grass sorghum and identifies important alleles of SbPRR37 that are critical for the utilization of this tropical grass in temperate zone grain and bioenergy production.

Functional annotation of the transcriptome of Sorghum bicolor in response to osmotic stress and abscisic acid.

BackgroundHigher plants exhibit remarkable phenotypic plasticity allowing them to adapt to an extensive range of environmental conditions. Sorghum is a cereal crop that exhibits exceptional tolerance to adverse conditions, in particular, water-limiting environments. This study utilized next generation sequencing (NGS) technology to examine the transcriptome of sorghum plants challenged with osmotic stress and exogenous abscisic acid (ABA) in order to elucidate genes and gene networks that contribute to sorghums tolerance to water-limiting environments with a long-term aim of developing strategies to improve plant productivity under drought.ResultsRNA-Seq results revealed transcriptional activity of 28,335 unique genes from sorghum root and shoot tissues subjected to polyethylene glycol (PEG)-induced osmotic stress or exogenous ABA. Differential gene expression analyses in response to osmotic stress and ABA revealed a strong interplay among various metabolic pathways including abscisic acid and 13-lipoxygenase, salicylic acid, jasmonic acid, and plant defense pathways. Transcription factor analysis indicated that groups of genes may be co-regulated by similar regulatory sequences to which the expressed transcription factors bind. We successfully exploited the data presented here in conjunction with published transcriptome analyses for rice, maize, and Arabidopsis to discover more than 50 differentially expressed, drought-responsive gene orthologs for which no function had been previously ascribed.ConclusionsThe present study provides an initial assemblage of sorghum genes and gene networks regulated by osmotic stress and hormonal treatment. We are providing an RNA-Seq data set and an initial collection of transcription factors, which offer a preliminary look into the cascade of global gene expression patterns that arise in a drought tolerant crop subjected to abiotic stress. These resources will allow scientists to query gene expression and functional annotation in response to drought.

Alignment of genetic maps and QTLs between inter- and intra-specific sorghum populations

To increase the value of associated molecular tools and also to begin to explore the degree to which interspecific and intraspecific genetic variation in Sorghum is attributable to corresponding genetic loci, we have aligned genetic maps derived from two sorghum populations that share one common parent (Sorghum bicolor L. Moench accession BTx623) but differ in morphological and evolutionarily distant alternate parents (S. propinquum or S. bicolor accession IS3620C). A total of 106 well-distributed DNA markers provide for map alignment, revealing only six nominal differences in marker order that are readily explained by sampling variation or mapping of paralogous loci. We also report a total of 61 new QTLs detected from 17 traits in these crosses. Among eight corresponding traits (some new, some previously published) that could be directly compared between the two maps, QTLs for two (tiller height and tiller number) were found to correspond in a non-random manner (P<0.05). For several other traits, correspondence of subsets of QTLs narrowly missed statistical significance. In particular, several QTLs for leaf senescence were near loci previously mapped for ‘stay-green’ that have been implicated by others in drought tolerance. These data provide strong validation for the value of molecular tools developed in the interspecific cross for utilization in cultivated sorghum, and begin to separate QTLs that distinguish among Sorghum species from those that are informative within the cultigen (S. bicolor).

Comprehensive Molecular Cytogenetic Analysis of Sorghum Genome Architecture: Distribution of Euchromatin, Heterochromatin, Genes and Recombination in Comparison to Rice

Cytogenetic maps of sorghum chromosomes 3–7, 9, and 10 were constructed on the basis of the fluorescence in situ hybridization (FISH) of ∼18–30 BAC probes mapped across each of these chromosomes. Distal regions of euchromatin and pericentromeric regions of heterochromatin were delimited for all 10 sorghum chromosomes and their DNA content quantified. Euchromatic DNA spans ∼50% of the sorghum genome, ranging from ∼60% of chromosome 1 (SBI-01) to ∼33% of chromosome 7 (SBI-07). This portion of the sorghum genome is predicted to encode ∼70% of the sorghum genes (∼1 gene model/12.3 kbp), assuming that rice and sorghum encode a similar number of genes. Heterochromatin spans ∼411 Mbp of the sorghum genome, a region characterized by a ∼34-fold lower rate of recombination and ∼3-fold lower gene density compared to euchromatic DNA. The sorghum and rice genomes exhibit a high degree of macrocolinearity; however, the sorghum genome is ∼2-fold larger than the rice genome. The distal euchromatic regions of sorghum chromosomes 3–7 and 10 are ∼1.8-fold larger overall and exhibit an ∼1.5-fold lower average rate of recombination than the colinear regions of the homeologous rice chromosomes. By contrast, the pericentromeric heterochromatic regions of these chromosomes are on average ∼3.6-fold larger in sorghum and recombination is suppressed ∼15-fold compared to the colinear regions of rice chromosomes.

Identification of genomic regions that affect grain-mould incidence and other traits of agronomic importance in sorghum

Abstract Grain-mould is a major problem in grain sorghum utilization as mouldy grain has a reduced quality due to the deterioration of the endosperm and reduced embryo viability. Here, our objective was to use genome mapping to improve knowledge of genetic variation and co-variation for grain-mould incidence and other inter-related agronomic traits. Grain-mould incidence, kernel-milling hardness, grain density, plant height, panicle peduncle length, foliar-disease incidence, and plant color were measured on 125 F5 genotypes derived from a cross of Sureño and RTx430. Quantitative trait loci were detected by means of 130 mapped markers (44 microsatellites, 85 AFLPs, one morphological-trait locus) distributed among ten linkage groups covering 970 cM. One to five QTLs affected each trait, with the exception of grain density for which no QTLs were detected. Grain-mould incidence was affected by five QTLs each accounting for between 10 and 23% of the phenotypic variance. The effects and relative positions of QTLs for grain-mould incidence were in accordance with the QTL distribution of several inter-related agronomic traits (e.g., plant height, peduncle length) and with the correlation between these phenotypic traits and grain-mould incidence. The detection of QTLs for grain-mould incidence was dependent on the environment, which is consistent with heritibility estimates that show strong environmental and genotype × environment effects. Several genomic regions affected multiple traits including one region that affected grain-mould incidence, plant height, panicle peduncle length, and grain-milling hardness, and a second region that influenced grain-mould (in four environments) and plant height. One genomic region, which harbors loci for plant color, influenced the severity of foliar disease symptoms and the incidence of grain-mould in one environment. Collectively QTLs detected in the present population explained between 10% and 55% of the phenotypic variance observed for a given trait.

Drought adaptation of stay-green sorghum is associated with canopy development, leaf anatomy, root growth, and water uptake

Summary The positive effects of stay-green quantitative trait loci on grain yield of sorghum under post-anthesis drought are emergent consequences of their effects on water-use patterns, resulting from changes in pre-anthesis canopy size.

Inheritance of inflorescence architecture in sorghum.

The grass inflorescence is the primary food source for humanity, and has been repeatedly shaped by human selection during the domestication of different cereal crops. Of all major cultivated cereals, sorghum [Sorghum bicolor (L.) Moench] shows the most striking variation in inflorescence architecture traits such as branch number and branch length, but the genetic basis of this variation is little understood. To study the inheritance of inflorescence architecture in sorghum, 119 recombinant inbred lines from an elite by exotic cross were grown in three environments and measured for 15 traits, including primary, secondary, and tertiary inflorescence branching. Eight characterized genes that are known to control inflorescence architecture in maize (Zea mays L.) and other grasses were mapped in sorghum. Two of these candidate genes, Dw3 and the sorghum ortholog of ramosa2, co-localized precisely with QTL of large effect for relevant traits. These results demonstrate the feasibility of using genomic and mutant resources from maize and rice (Oryza sativa L.) to investigate the inheritance of complex traits in related cereals.

Large Intraspecific Haplotype Variability at the Rph7 Locus Results from Rapid and Recent Divergence in the Barley Genome

To study genome evolution and diversity in barley (Hordeum vulgare), we have sequenced and compared more than 300 kb of sequence spanning the Rph7 leaf rust disease resistance gene in two barley cultivars. Colinearity was restricted to five genic and two intergenic regions representing <35% of the two sequences. In each interval separating the seven conserved regions, the number and type of repetitive elements were completely different between the two homologous sequences, and a single gene was absent in one cultivar. In both cultivars, the nonconserved regions consisted of ∼53% repetitive sequences mainly represented by long-terminal repeat retrotransposons that have inserted <1 million years ago. PCR-based analysis of intergenic regions at the Rph7 locus and at three other independent loci in 41 H. vulgare lines indicated large haplotype variability in the cultivated barley gene pool. Together, our data indicate rapid and recent divergence at homologous loci in the genome of H. vulgare, possibly providing the molecular mechanism for the generation of high diversity in the barley gene pool. Finally, comparative analysis of the gene composition in barley, wheat (Triticum aestivum), rice (Oryza sativa), and sorghum (Sorghum bicolor) suggested massive gene movements at the Rph7 locus in the Triticeae lineage.