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Pan-genome Analysis in Sorghum Highlights the Extent of Genomic Variation and Sugarcane Aphid Resistance Genes


First published on 4 January 2021

Bo Wang, Yinping Jiao, Kapeel Chougule, Andrew Olson, Jian Huang, Victor Llaca, Kevin Fengler, Xuehong Wei, Liya Wang, Xiaofei Wang, Michael Regulski, Jorg Drenkow, Thomas Gingeras, Chad Hayes, J. Scott Armstrong, Yinghua Huang, Zhanguo Xin, Doreen Ware

Sorghum bicolor, one of the most important grass crops around the world, harbors a high degree of genetic diversity. We constructed chromosome-level genome assemblies for two important sorghum inbred lines, Tx2783 and RTx436. The final high-quality reference assemblies consist of 19 and 18 scaffolds, respectively, with contig N50 values of 25.6 and 20.3 Mb. Genes were annotated using evidence-based and de novo gene predictors, and RAMPAGE data demonstrate that transcription start sites were effectively captured. Together with other public sorghum genomes, BTx623, RTx430, and Rio, extensive structural variations (SVs) of various sizes were characterized using Tx2783 as a reference. Genome-wide scanning for disease resistance (R) genes revealed high levels of diversity among these five sorghum accessions. To characterize sugarcane aphid (SCA) resistance in Tx2783, we mapped the resistance region on chromosome 6 using a recombinant inbred line (RIL) population and found a SV of 191 kb containing a cluster of R genes in Tx2783. Using Tx2783 as a backbone, along with the SVs, we constructed a pan-genome to support alignment of resequencing data from 62 sorghum accessions, and then identified core and dispensable genes using this population. This study provides the first overview of the extent of genomic structural variations and R genes in the sorghum population, and reveals potential targets for breeding of SCA resistance.

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BSAseq: an interactive and integrated web-based workflow for identification of causal mutations in bulked F2 populations


First published on 11 August 2020

Pubmed ID: 32777814

Keywords: No keywords in Pubmed

Wang L, Lu Z, Regulski M, Jiao Y, Chen J, Ware D, Xin Z

SUMMARY: With the advance of next-generation sequencing (NGS) technologies and reductions in the costs of these techniques, bulked segregant analysis (BSA) has become not only a powerful tool for mapping quantitative trait loci (QTL) but also a useful way to identify causal gene mutations underlying phenotypes of interest. However, due to the presence of background mutations and errors in sequencing, genotyping, and reference assembly, it is often difficult to distinguish true causal mutations from background mutations. In this study, we developed the BSAseq workflow, which includes an automated bioinformatics analysis pipeline with a probabilistic model for estimating the linked region (the region linked to the causal mutation) and an interactive Shiny web application for visualizing the results. We deeply sequenced a sorghum male-sterile parental line (ms8) to capture the majority of background mutations in our bulked F2 data. We applied the workflow to 11 bulked sorghum F2 populations and 1 rice F2 population and identified the true causal mutation in each population. The workflow is intuitive and straightforward, facilitating its adoption by users without bioinformatics analysis skills. We anticipate that the BSAseq workflow will be broadly applicable to the identification of causal mutations for many phenotypes of interest.AVAILABILITY: BSAseq is freely available on https://www.sciapps.org/page/bsa.SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.

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Genome-wide association study reveals that different pathways contribute to grain quality variation in sorghum (Sorghum bicolor).


First published on 2 February 2020

Pubmed ID: 32005168

Keywords: Amino acids, Genome-wide association study, Grain quality, Sorghum, Starch, Tannins

Kimani W, Zhang LM, Wu XY, Hao HQ, Jing HC

BACKGROUND: In sorghum (Sorghum bicolor), one paramount breeding objective is to increase grain quality. The nutritional quality and end use value of sorghum grains are primarily influenced by the proportions of tannins, starch and proteins, but the genetic basis of these grain quality traits remains largely unknown. This study aimed to dissect the natural variation of sorghum grain quality traits and identify the underpinning genetic loci by genome-wide association study.RESULTS: Levels of starch, tannins and 17 amino acids were quantified in 196 diverse sorghum inbred lines, and 44 traits based on known metabolic pathways and biochemical interactions amongst the 17 amino acids calculated. A Genome-wide association study (GWAS) with 3,512,517 SNPs from re-sequencing data identified 14, 15 and 711 significant SNPs which represented 14, 14, 492 genetic loci associated with levels of tannins, starch and amino acids in sorghum grains, respectively. Amongst these significant SNPs, two SNPs were associated with tannin content on chromosome 4 and colocalized with three previously identified loci for Tannin1, and orthologs of Zm1 and TT16 genes. One SNP associated with starch content colocalized with sucrose phosphate synthase gene. Furthermore, homologues of opaque1 and opaque2 genes associated with amino acid content were identified. Using the KEGG pathway database, six and three candidate genes of tannins and starch were mapped into 12 and 3 metabolism pathways, respectively. Thirty-four candidate genes were mapped into 16 biosynthetic and catabolic pathways of amino acids. We finally reconstructed the biosynthetic pathways for aspartate and branched-chain amino acids based on 15 candidate genes identified in this study.CONCLUSION: Promising candidate genes associated with grain quality traits have been identified in the present study. Some of them colocalized with previously identified genetic regions, but novel candidate genes involved in various metabolic pathways which influence grain quality traits have been dissected. Our study acts as an entry point for further validation studies to elucidate the complex mechanisms controlling grain quality traits such as tannins, starch and amino acids in sorghum.

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Evaluation of genetic diversity, agronomic traits, and anthracnose resistance in the NPGS Sudan Sorghum Core collection


First published on 30 January 2020

Pubmed ID: 31992189

Keywords: Anthracnose, Genome-wide association analysis, Genotyping-by-sequencing, NPGS sorghum germplasm, Population structure

Cuevas HE, Prom LK

BACKGROUND: The United States Department of Agriculture (USDA) National Plant Germplasm System (NPGS) sorghum core collection contains 3011 accessions randomly selected from 77 countries. Genomic and phenotypic characterization of this core collection is necessary to encourage and facilitate its utilization in breeding programs and to improve conservation efforts. In this study, we examined the genome sequences of 318 accessions belonging to the NPGS Sudan sorghum core set, and characterized their agronomic traits and anthracnose resistance response.RESULTS: We identified 183,144 single nucleotide polymorphisms (SNPs) located within or in proximity of 25,124 annotated genes using the genotyping-by-sequencing (GBS) approach. The core collection was genetically highly diverse, with an average pairwise genetic distance of 0.76 among accessions. Population structure and cluster analysis revealed five ancestral populations within the Sudan core set, with moderate to high level of genetic differentiation. In total, 171 accessions (54%) were assigned to one of these populations, which covered 96% of the total genomic variation. Genome scan based on Tajima's D values revealed two populations under balancing selection. Phenotypic analysis showed differences in agronomic traits among the populations, suggesting that these populations belong to different ecogeographical regions. A total of 55 accessions were resistant to anthracnose; these accessions could represent multiple resistance sources. Genome-wide association study based on fixed and random model Circulating Probability (farmCPU) identified genomic regions associated with plant height, flowering time, panicle length and diameter, and anthracnose resistance response. Integrated analysis of the Sudan core set and sorghum association panel indicated that a large portion of the genetic variation in the Sudan core set might be present in breeding programs but remains unexploited within some clusters of accessions.CONCLUSIONS: The NPGS Sudan core collection comprises genetically and phenotypically diverse germplasm with multiple anthracnose resistance sources. Population genomic analysis could be used to improve screening efforts and identify the most valuable germplasm for breeding programs. The new GBS data set generated in this study represents a novel genomic resource for plant breeders interested in mining the genetic diversity of the NPGS sorghum collection.

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Fungal community assembly in drought-stressed sorghum shows stochasticity, selection, and universal ecological dynamics


First published on 9 January 2020

Pubmed ID: 31911594

Keywords: No keywords in Pubmed

Gao C, Montoya L, Xu L, Madera M, Hollingsworth J, Purdom E, Singan V, Vogel J, Hutmacher RB, Dahlberg JA, Coleman-Derr D, Lemaux PG, Taylor JW

Community assembly of crop-associated fungi is thought to be strongly influenced by deterministic selection exerted by the plant host, rather than stochastic processes. Here we use a simple, sorghum system with abundant sampling to show that stochastic forces (drift or stochastic dispersal) act on fungal community assembly in leaves and roots early in host development and when sorghum is drought stressed, conditions when mycobiomes are small. Unexpectedly, we find no signal for stochasticity when drought stress is relieved, likely due to renewed selection by the host. In our experimental system, the host compartment exerts the strongest effects on mycobiome assembly, followed by the timing of plant development and lastly by plant genotype. Using a dissimilarity-overlap approach, we find a universality in the forces of community assembly of the mycobiomes of the different sorghum compartments and in functional guilds of fungi.

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Transcriptomic analysis of field-droughted sorghum from seedling to maturity reveals biotic and metabolic responses


First published on 7 December 2019

Pubmed ID: 31806758

Keywords: RNA-Seq, S. bicolor, arbuscular mycorrhizal fungi, drought

Varoquaux N, Cole B, Gao C, Pierroz G, Baker CR, Patel D, Madera M, Jeffers T, Hollingsworth J, Sievert J, Yoshinaga Y, Owiti JA, Singan VR, DeGraaf S, Xu L, Blow MJ, Harrison MJ, Visel A, Jansson C, Niyogi KK, Hutmacher R, Coleman-Derr D, O'Malley RC, Taylor JW, Dahlberg J, Vogel JP, Lemaux PG, Purdom E

Drought is the most important environmental stress limiting crop yields. The C4 cereal sorghum [Sorghum bicolor (L.) Moench] is a critical food, forage, and emerging bioenergy crop that is notably drought-tolerant. We conducted a large-scale field experiment, imposing preflowering and postflowering drought stress on 2 genotypes of sorghum across a tightly resolved time series, from plant emergence to postanthesis, resulting in a dataset of nearly 400 transcriptomes. We observed a fast and global transcriptomic response in leaf and root tissues with clear temporal patterns, including modulation of well-known drought pathways. We also identified genotypic differences in core photosynthesis and reactive oxygen species scavenging pathways, highlighting possible mechanisms of drought tolerance and of the delayed senescence, characteristic of the stay-green phenotype. Finally, we discovered a large-scale depletion in the expression of genes critical to arbuscular mycorrhizal (AM) symbiosis, with a corresponding drop in AM fungal mass in the plants' roots.

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The Impact of Sorghum Growth Stage and Resistance on Life History of Sugarcane Aphids (Hemiptera: Aphididae).


First published on 22 November 2019

Pubmed ID: 31751451

Keywords: fecundity, host plant growth stage, host plant resistance, intrinsic rate of increase, population doubling

Neupane SB, Kerns DL, Szczepaniec A

Recent invasion of a new haplotype of sugarcane aphid (Melanaphis sacchari Zehntner) resulted in severe outbreaks of the aphids in sorghum (Sorghum bicolor L. Moench [Poales: Poaceae]) across the United States. Previous research and field observations suggested that both sorghum resistance and growth stage were important to their population dynamics in the field and hence effective and timely management to minimize economic losses associated with this pest. To explore this, we designed greenhouse experiments to quantify fecundity, prereproductive period, and longevity of sugarcane aphids across several vegetative and reproductive growth stages of a resistant (DKS 37-07) and susceptible (M77GB52 and DKS 38-88) sorghum hybrids commonly used in sorghum production. We found significant effects of sorghum resistance and growth stage on several life history traits and demographics parameters of sugarcane aphids. We did not, however, note any significant interactive effects of resistance and phenology on any of the response variables. Sugarcane aphids exposed to the susceptible sorghum produced significantly more offspring, had significantly greater intrinsic and finite rates of increase, and significantly shorter population doubling time than the aphids feeding on resistant sorghum. On the other hand, the impact of sorghum growth stage had more variable effects on life history of sugarcane aphids that were most frequently evident at the late reproductive stages. These outcomes support our hypothesis that sorghum in late stages of growth tends to be a better host for sugarcane aphids and highlight the importance of sorghum growth stage to sugarcane aphid life history and population growth potential.

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Sorghum MSD3 Encodes an ω-3 Fatty Acid Desaturase that Increases Grain Number by Reducing Jasmonic Acid Levels


First published on 31 October 2019

Pubmed ID: 31661847

Keywords: MutMap, fatty acid desaturase, grain number, jasmonic acid, msd, multiseeded, sorghum

Dampanaboina L, Jiao Y, Chen J, Gladman N, Chopra R, Burow G, Hayes C, Christensen SA, Burke J, Ware D, Xin Z

Grain number per panicle is an important component of grain yield in sorghum (Sorghum bicolor (L.)) and other cereal crops. Previously, we reported that mutations in multi-seeded 1 (MSD1) and MSD2 genes result in a two-fold increase in grain number per panicle due to the restoration of the fertility of the pedicellate spikelets, which invariably abort in natural sorghum accessions. Here, we report the identification of another gene, MSD3, which is also involved in the regulation of grain numbers in sorghum. Four bulked F2 populations from crosses between BTx623 and each of the independent msd mutants p6, p14, p21, and p24 were sequenced to 20× coverage of the whole genome on a HiSeq 2000 system. Bioinformatic analyses of the sequence data showed that one gene, Sorbi_3001G407600, harbored homozygous mutations in all four populations. This gene encodes a plastidial ω-3 fatty acid desaturase that catalyzes the conversion of linoleic acid (18:2) to linolenic acid (18:3), a substrate for jasmonic acid (JA) biosynthesis. The msd3 mutants had reduced levels of linolenic acid in both leaves and developing panicles that in turn decreased the levels of JA. Furthermore, the msd3 panicle phenotype was reversed by treatment with methyl-JA (MeJA). Our characterization of MSD1, MSD2, and now MSD3 demonstrates that JA-regulated processes are critical to the msd phenotype. The identification of the MSD3 gene reveals a new target that could be manipulated to increase grain number per panicle in sorghum, and potentially other cereal crops, through the genomic editing of MSD3 functional orthologs.

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Large-scale GWAS in sorghum reveals common genetic control of grain size among cereals


First published on 30 October 2019

Pubmed ID: 31659829

Keywords: GWAS, QTL, cereal crops, grain size, orthologues, sorghum

Tao Y, Zhao X, Wang X, Hathorn A, Hunt C, W Cruickshank A, van Oosterom EJ, Godwin ID, Mace ES, Jordan DR

Grain size is a key yield component of cereal crops and a major quality attribute. It is determined by a genotype's genetic potential and its capacity to fill the grains. This study aims to dissect the genetic architecture of grain size in sorghum. An integrated genome wide association study (GWAS) was conducted using a diversity panel (n=837) and a BC-NAM population (n=1,421). To isolate genetic effects associated with genetic potential of grain size, rather than the genotype's capacity to fill the grains, a treatment of removing half of the panicle was imposed during flowering. Extensive and highly heritable variation in grain size was observed in both populations in 5 field trials and 81 grain size QTL were identified in subsequent GWAS. These QTL were enriched for orthologues of known grain size genes in rice and maize, and had significant overlap with SNPs associated with grain size in rice and maize, supporting common genetic control of this trait among cereals. Grain size genes with opposite effect on grain number were less likely to overlap with the grain size QTL from this study, indicating the treatment facilitated identification of genetic regions related to the genetic potential of grain size. These results enhance understanding of the genetic architecture of grain size in cereal, and pave the way for exploration of underlying molecular mechanisms and manipulation of this trait in breeding practices.

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Genetic Architecture of Chilling Tolerance in Sorghum Dissected with a Nested Association Mapping Population


First published on 16 October 2019

Pubmed ID: 31611346

Keywords: antagonistic pleiotropy, climate adaptation, cold tolerance, crop evolution, linkage drag, multiparental population

Marla SR, Burow G, Chopra R, Hayes C, Olatoye MO, Felderhoff T, Hu Z, Raymundo R, Perumal R, Morris GP

Dissecting the genetic architecture of stress tolerance in crops is critical to understand and improve adaptation. In temperate climates, early planting of chilling-tolerant varieties could provide longer growing seasons and drought escape, but chilling tolerance (75,000 data points from ∼16,000 plots) in multi-environment field trials in Kansas under natural chilling stress (sown 30-45 days early) and normal growing conditions. Joint linkage mapping with early-planted field phenotypes revealed an oligogenic architecture, with 5-10 chilling tolerance loci explaining 20-41% of variation. Surprisingly, several of the major chilling tolerance loci colocalize precisely with the classical grain tannin (Tan1 and Tan2) and dwarfing genes (Dw1 and Dw3) that were under strong directional selection in the US during the 20th century. These findings suggest that chilling sensitivity was inadvertently selected due to coinheritance with desired nontannin and dwarfing alleles. The characterization of genetic architecture with NAM reveals why past chilling tolerance breeding was stymied and provides a path for genomicsenabled breeding of chilling tolerance.

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Fertility of Pedicellate Spikelets in Sorghum Is Controlled by a Jasmonic Acid Regulatory Module


First published on 11 October 2019

Pubmed ID: 31597271

Keywords: gene expression, jasmonic acid signaling, plant development, transcriptional regulators

Gladman N, Jiao Y, Lee YK, Zhang L, Chopra R, Regulski M, Burow G, Hayes C, Christensen SA, Dampanaboina L, Chen J, Burke J, Ware D, Xin Z

As in other cereal crops, the panicles of sorghum (Sorghum bicolor (L.) Moench) comprise two types of floral spikelets (grass flowers). Only sessile spikelets (SSs) are capable of producing viable grains, whereas pedicellate spikelets (PSs) cease development after initiation and eventually abort. Consequently, grain number per panicle (GNP) is lower than the total number of flowers produced per panicle. The mechanism underlying this differential fertility is not well understood. To investigate this issue, we isolated a series of ethyl methane sulfonate (EMS)-induced multiseeded (msd) mutants that result in full spikelet fertility, effectively doubling GNP. Previously, we showed that MSD1 is a TCP (Teosinte branched/Cycloidea/PCF) transcription factor that regulates jasmonic acid (JA) biosynthesis, and ultimately floral sex organ development. Here, we show that MSD2 encodes a lipoxygenase (LOX) that catalyzes the first committed step of JA biosynthesis. Further, we demonstrate that MSD1 binds to the promoters of MSD2 and other JA pathway genes. Together, these results show that a JA-induced module regulates sorghum panicle development and spikelet fertility. The findings advance our understanding of inflorescence development and could lead to new strategies for increasing GNP and grain yield in sorghum and other cereal crops.

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Reconstructing the Transcriptional Ontogeny of Maize and Sorghum Supports an Inverse Hourglass Model of Inflorescence Development


First published on 8 October 2019

Pubmed ID: 31587998

Keywords: comparative transcriptomics, developmental hourglass, evo-devo, evolution of development, maize genetics, phylostratigraphy, sorghum inflorescence

Leiboff S, Hake S

Assembling meaningful comparisons between species is a major limitation in studying the evolution of organismal form. To understand development in maize and sorghum, closely related species with architecturally distinct inflorescences, we collected RNA-seq profiles encompassing inflorescence body-plan specification in both species. We reconstructed molecular ontogenies from 40 B73 maize tassels and 47 BTx623 sorghum panicles and separated them into transcriptional stages. To discover new markers of inflorescence development, we used random forest machine learning to determine stage by RNA-seq. We used two descriptions of transcriptional conservation to identify hourglass-like stages during inflorescence development. Despite a relatively short 12 million years since their last common ancestor, we found maize and sorghum inflorescences are most different during their hourglass-like stages of development, following an inverse-hourglass model of development. We discuss whether agricultural selection may account for the rapid divergence signatures in these species and the observed separation of evolutionary pressure and developmental reprogramming.

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Genome-wide identification and expression profile analysis of nuclear factor Y family genes in Sorghum bicolor L. (Moench).


First published on 20 September 2019

Pubmed ID: 31536532

Keywords: No keywords in Pubmed

Maheshwari P, Kummari D, Palakolanu SR, Nagasai Tejaswi U, Nagaraju M, Rajasheker G, Jawahar G, Jalaja N, Rathnagiri P, Kavi Kishor PB

Members of the plant Heme Activator Protein (HAP) or NUCLEAR FACTOR Y (NF-Y) are trimeric transcription factor complexes composed of the NF-YA, NF-YB and NF-YC subfamilies. They bind to the CCAAT box in the promoter regions of the target genes and regulate gene expressions. Plant NF-Ys were reported to be involved in adaptation to several abiotic stresses as well as in development. In silico analysis of Sorghum bicolor genome resulted in the identification of a total of 42 NF-Y genes, among which 8 code for the SbNF-YA, 19 for SbNF-YB and 15 for the SbNF-YC subunits. Analysis was also performed to characterize gene structures, chromosomal distribution, duplication status, protein subcellular localizations, conserved motifs, ancestral protein sequences, miRNAs and phylogenetic tree construction. Phylogenetic relationships and ortholog predictions displayed that sorghum has additional NF-YB genes with unknown functions in comparison with Arabidopsis. Analysis of promoters revealed that they harbour many stress-related cis-elements like ABRE and HSE, but surprisingly, DRE and MYB elements were not detected in any of the subfamilies. SbNF-YA1, 2, and 6 were found upregulated under 200 mM salt and 200 mM mannitol stresses. While NF-YA7 appeared associated with high temperature (40°C) stress, NF-YA8 was triggered by both cold (4°C) and high temperature stresses. Among NF-YB genes, 7, 12, 15, and 16 were induced under multiple stress conditions such as salt, mannitol, ABA, cold and high temperatures. Likewise, NF-YC 6, 11, 12, 14, and 15 were enhanced significantly in a tissue specific manner under multiple abiotic stress conditions. Majority of the mannitol (drought)-inducible genes were also induced by salt, high temperature stresses and ABA. Few of the high temperature stress-induced genes are also induced by cold stress (NF-YA2, 4, 6, 8, NF-YB2, 7, 10, 11, 12, 14, 16, 17, NF-YC4, 6, 12, and 13) thus suggesting a cross talk among them. This work paves the way for investigating the roles of diverse sorghum NF-Y proteins during abiotic stress responses and provides an insight into the evolution of diverse NF-Y members.

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Response of seedling growth and physiology of Sorghum bicolor (L.) Moench to saline-alkali stress


First published on 31 July 2019

Pubmed ID: 31361760

Keywords: No keywords in Pubmed

Sun J, He L, Li T

Soil salinization is a serious problem that affects the seedling growth in many regions. A greenhouse experiment was carried to investigate the adaptation ability of seedlings (Sorghum bicolor (L.) Moench.) in coastal saline alkaline environment. Seedlings of sorghum were treated by different salt and alkali stress (NaCl: Na2SO4: NaHCO3 were 2:1:0, 2:1:1, 2:1:2). The treatments consisted of three levels of salinity (100, 200 and 300 mmol/L) and pH values were 7.08, 8.78 and 9.04. The results showed that the seedlings of sorghum have good adaptability to salt stress under low pH (pH ≤7.08). The plant height, the maximum leave areas of seedlings all dropped and root length first ascended and then descended with the increasing of salt and alkali stress. The contents of Chlorophyll b degraded significantly under salt and alkali stress. Salt and alkali stress stimulated the accumulation of organic solutes (proline and protein) and inorganic ions (Na+, Cl-, SO42-). Our results showed that salt and alkali stress have significant effect on growth indexes except root length and the interaction effect has significantly on physiology.

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Transcriptome analysis reveals candidate genes related to phosphorus starvation tolerance in sorghum


First published on 13 July 2019

Pubmed ID: 31296169

Keywords: Candidate genes, Malate, Phosphorus starvation, Root development, Sorghum, Transcriptome analysis

Zhang J, Jiang F, Shen Y, Zhan Q, Bai B, Chen W, Chi Y

BACKGROUND: Phosphorus (P) deficiency in soil is a worldwide issue and a major constraint on the production of sorghum, which is an important staple food, forage and energy crop. The depletion of P reserves and the increasing price of P fertilizer make fertilizer application impractical, especially in developing countries. Therefore, identifying sorghum accessions with low-P tolerance and understanding the underlying molecular basis for this tolerance will facilitate the breeding of P-efficient plants, thereby resolving the P crisis in sorghum farming. However, knowledge in these areas is very limited.RESULTS: The 29 sorghum accessions used in this study demonstrated great variability in their tolerance to low-P stress. The internal P content in the shoot was correlated with P tolerance. A low-P-tolerant accession and a low-P-sensitive accession were chosen for RNA-seq analysis to identify potential underlying molecular mechanisms. A total of 2089 candidate genes related to P starvation tolerance were revealed and found to be enriched in 11 pathways. Gene Ontology (GO) enrichment analyses showed that the candidate genes were associated with oxidoreductase activity. In addition, further study showed that malate affected the length of the primary root and the number of tips in sorghum suffering from low-P stress.CONCLUSIONS: Our results show that acquisition of P from soil contributes to low-P tolerance in different sorghum accessions; however, the underlying molecular mechanism is complicated. Plant hormone (including auxin, ethylene, jasmonic acid, salicylic acid and abscisic acid) signal transduction related genes and many transcriptional factors were found to be involved in low-P tolerance in sorghum. The identified accessions will be useful for breeding new sorghum varieties with enhanced P starvation tolerance.

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Metabolomics of sorghum roots during nitrogen stress reveals compromised metabolic capacity for salicylic acid biosynthesis


First published on 28 June 2019

Pubmed ID: 31245765

Keywords: metabolism, metabolomics, microbiome, nitrogen, rhizosphere, roots, salicylic acid, sorghum, stress

Sheflin AM, Chiniquy D, Yuan C, Goren E, Kumar I, Braud M, Brutnell T, Eveland AL, Tringe S, Liu P, Kresovich S, Marsh EL, Schachtman DP, Prenni JE

Sorghum (Sorghum bicolor [L.] Moench) is the fifth most productive cereal crop worldwide with some hybrids having high biomass yield traits making it promising for sustainable, economical biofuel production. To maximize biofuel feedstock yields, a more complete understanding of metabolic responses to low nitrogen (N) will be useful for incorporation in crop improvement efforts. In this study, 10 diverse sorghum entries (including inbreds and hybrids) were field-grown under low and full N conditions and roots were sampled at two time points for metabolomics and 16S amplicon sequencing. Roots of plants grown under low N showed altered metabolic profiles at both sampling dates including metabolites important in N storage and synthesis of aromatic amino acids. Complementary investigation of the rhizosphere microbiome revealed dominance by a single operational taxonomic unit (OTU) in an early sampling that was taxonomically assigned to the genus Pseudomonas. Abundance of this Pseudomonas OTU was significantly greater under low N in July and was decreased dramatically in September. Correlation of Pseudomonas abundance with root metabolites revealed a strong negative association with the defense hormone salicylic acid (SA) under full N but not under low N, suggesting reduced defense response. Roots from plants with N stress also contained reduced phenylalanine, a precursor for SA, providing further evidence for compromised metabolic capacity for defense response under low N conditions. Our findings suggest that interactions between biotic and abiotic stresses may affect metabolic capacity for plant defense and need to be concurrently prioritized as breeding programs become established for biofuels production on marginal soils.

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A new reference genome for Sorghum bicolor reveals high levels of sequence similarity between sweet and grain genotypes: implications for the genetics of sugar metabolism


First published on 28 May 2019

Pubmed ID: 31133004

Keywords: Gene expression, Genomics, Sorghum, Sugar metabolism, Sugar transport

Cooper EA, Brenton ZW, Flinn BS, Jenkins J, Shu S, Flowers D, Luo F, Wang Y, Xia P, Barry K, Daum C, Lipzen A, Yoshinaga Y, Schmutz J, Saski C, Vermerris W, Kresovich S

BACKGROUND: The process of crop domestication often consists of two stages: initial domestication, where the wild species is first cultivated by humans, followed by diversification, when the domesticated species are subsequently adapted to more environments and specialized uses. Selective pressure to increase sugar accumulation in certain varieties of the cereal crop Sorghum bicolor is an excellent example of the latter; this has resulted in pronounced phenotypic divergence between sweet and grain-type sorghums, but the genetic mechanisms underlying these differences remain poorly understood.RESULTS: Here we present a new reference genome based on an archetypal sweet sorghum line and compare it to the current grain sorghum reference, revealing a high rate of nonsynonymous and potential loss of function mutations, but few changes in gene content or overall genome structure. We also use comparative transcriptomics to highlight changes in gene expression correlated with high stalk sugar content and show that changes in the activity and possibly localization of transporters, along with the timing of sugar metabolism play a critical role in the sweet phenotype.CONCLUSIONS: The high level of genomic similarity between sweet and grain sorghum reflects their historical relatedness, rather than their current phenotypic differences, but we find key changes in signaling molecules and transcriptional regulators that represent new candidates for understanding and improving sugar metabolism in this important crop.

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Global Responses of Resistant and Susceptible Sorghum (Sorghum bicolor) to Sugarcane Aphid (Melanaphis sacchari)


First published on 12 March 2019

Pubmed ID: 30853964

Keywords: Melanaphis sacchari, Sorghum bicolor, plant–insect interactions, sorghum, sugarcane aphid

Tetreault HM, Grover S, Scully ED, Gries T, Palmer NA, Sarath G, Louis J, Sattler SE

The sugarcane aphid (Melanaphis sacchari) has emerged as a significant pest for sorghum. The use of sugarcane aphid-resistant sorghum germplasm with integrated pest management strategies appears to be an excellent solution to this problem. In this study, a resistant line (RTx2783) and a susceptible line (A/BCK60) were used to characterize the differences in plant responses to the sugarcane aphid through a series of experiments, which examined global sorghum gene expression, aphid feeding behavior and inheritance of aphid resistance. The global transcriptomic responses to sugarcane aphids in resistant and susceptible plants were identified using RNA-seq and compared to the expression profiles of uninfested plants at 5, 10, and 15 days post-infestation. The expression of genes from several functional categories were altered in aphid-infested susceptible plants, which included genes related to cell wall modification, photosynthesis and phytohormone biosynthesis. In the resistant line, only 31 genes were differentially expressed in the infested plants relative to uninfested plants over the same timecourse. However, network analysis of these transcriptomes identified a co-expression module where the expression of multiple sugar and starch associated genes were repressed in infested resistant plants at 5 and 10 days. Several nucleotide-binding-site, leucine-rich repeat (NBS-LRR) and disease resistance genes similar to aphid resistance genes identified in other plants are identified in the current study which may be involved in sugarcane aphid resistance. The electrical penetration graph (EPG) results indicated that sugarcane aphid spent approximately twice as long in non-probing phase, and approximately a quarter of time in phloem ingestion phase on the resistant and F1 plants compared to susceptible plant. Additionally, network analysis identified a phloem protein 2 gene expressed in both susceptible and resistant plants early (day 5) of infestation, which may contribute to defense against aphid feeding within sieve elements. The resistant line RTx2783 displayed both antixenosis and antibiosis modes of resistance based on EPG and choice bioassays between susceptible, resistant and F1 plants. Aphid resistance from RTx2783 segregated as a single dominant locus in the F2 generation, which will enable breeders to rapidly develop sugarcane aphid-resistant hybrids using RTx2783 as the male parent.

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The genetic architecture of phosphorus efficiency in sorghum involves pleiotropic QTL for root morphology and grain yield under low phosphorus availability in the soil


First published on 2 March 2019

Pubmed ID: 30819116

Keywords: Acid soils, Phosphorus deficiency, Phosphorus stress, Root system architecture

Bernardino KC, Pastina MM, Menezes CB, de Sousa SM, Maciel LS, Carvalho G, Guimarães CT, Barros BA, da Costa E Silva L, Carneiro PCS, Schaffert RE, Kochian LV, Magalhaes JV

BACKGROUND: Phosphorus (P) fixation on aluminum (Al) and iron (Fe) oxides in soil clays restricts P availability for crops cultivated on highly weathered tropical soils, which are common in developing countries. Hence, P deficiency becomes a major obstacle for global food security. We used multi-trait quantitative trait loci (QTL) mapping to study the genetic architecture of P efficiency and to explore the importance of root traits on sorghum grain yield on a tropical low-P soil.RESULTS: P acquisition efficiency was the most important component of P efficiency, and both traits were highly correlated with grain yield under low P availability. Root surface area was positively associated with grain yield. The guinea parent, SC283, contributed 58% of all favorable alleles detected by single-trait mapping. Multi-trait mapping detected 14 grain yield and/or root morphology QTLs. Tightly linked or pleiotropic QTL underlying the surface area of fine roots (1-2 mm in diameter) and grain yield were detected at positions 1-7 megabase pairs (Mb) and 71 Mb on chromosome 3, respectively, and a root diameter/grain yield QTL was detected at 7 Mb on chromosome 7. All these QTLs were near sorghum homologs of the rice serine/threonine kinase, OsPSTOL1. The SbPSTOL1 genes on chromosome 3, Sb03g006765 at 7 Mb and Sb03g031690 at 60 Mb were more highly expressed in SC283, which donated the favorable alleles at all QTLs found nearby SbPSTOL1 genes. The Al tolerance gene, SbMATE, may also influence a grain yield QTL on chromosome 3. Another PSTOL1-like gene, Sb07g02840, appears to enhance grain yield via small increases in root diameter. Co-localization analyses suggested a role for other genes, such as a sorghum homolog of the Arabidopsis ubiquitin-conjugating E2 enzyme, phosphate 2 (PHO2), on grain yield advantage conferred by the elite parent, BR007 allele.CONCLUSIONS: Genetic determinants conferring higher root surface area and slight increases in fine root diameter may favor P uptake, thereby enhancing grain yield under low-P availability in the soil. Molecular markers for SbPSTOL1 genes and for QTL increasing grain yield by non-root morphology-based mechanisms hold promise in breeding strategies aimed at developing sorghum cultivars adapted to low-P soils.

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A chromosome-scale assembly of the sorghum genome using nanopore sequencing and optical mapping


First published on 20 November 2018

Pubmed ID: 30451840

Keywords: No keywords in Pubmed

Deschamps S, Zhang Y, Llaca V, Ye L, Sanyal A, King M, May G, Lin H

Long-read sequencing technologies have greatly facilitated assemblies of large eukaryotic genomes. In this paper, Oxford Nanopore sequences generated on a MinION sequencer are combined with Bionano Genomics Direct Label and Stain (DLS) optical maps to generate a chromosome-scale de novo assembly of the repeat-rich Sorghum bicolor Tx430 genome. The final assembly consists of 29 scaffolds, encompassing in most cases entire chromosome arms. It has a scaffold N50 of 33.28 Mbps and covers 90% of the expected genome length. A sequence accuracy of 99.85% is obtained after aligning the assembly against Illumina Tx430 data and 99.6% of the 34,211 public gene models align to the assembly. Comparisons of Tx430 and BTx623 DLS maps against the public BTx623 v3.0.1 genome assembly suggest substantial discrepancies whose origin remains to be determined. In summary, this study demonstrates that informative assemblies of complex plant genomes can be generated by combining nanopore sequencing with DLS optical maps.

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The Sorghum QTL Atlas: a powerful tool for trait dissection, comparative genomics and crop improvement


First published on 22 October 2018

Pubmed ID: 30343386

Keywords: No keywords in Pubmed

Mace E, Innes D, Hunt C, Wang X, Tao Y, Baxter J, Hassall M, Hathorn A, Jordan D

We describe the development and application of the Sorghum QTL Atlas, a high-resolution, open-access research platform to facilitate candidate gene identification across three cereal species, sorghum, maize and rice. The mechanisms governing the genetic control of many quantitative traits are only poorly understood and have yet to be fully exploited. Over the last two decades, over a thousand QTL and GWAS studies have been published in the major cereal crops including sorghum, maize and rice. A large body of information has been generated on the genetic basis of quantitative traits, their genomic location, allelic effects and epistatic interactions. However, such QTL information has not been widely applied by cereal improvement programs and genetic researchers worldwide. In part this is due to the heterogeneous nature of QTL studies which leads QTL reliability variation from study to study. Using approaches to adjust the QTL confidence interval, this platform provides access to the most updated sorghum QTL information than any database available, spanning 23 years of research since 1995. The QTL database provides information on the predicted gene models underlying the QTL CI, across all sorghum genome assembly gene sets and maize and rice genome assemblies and also provides information on the diversity of the underlying genes and information on signatures of selection in sorghum. The resulting high-resolution, open-access research platform facilitates candidate gene identification across 3 cereal species, sorghum, maize and rice. Using a number of trait examples, we demonstrate the power and resolution of the resource to facilitate comparative genomics approaches to provide a bridge between genomics and applied breeding.

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Isolation and Analysis of Microbial Communities in Soil, Rhizosphere, and Roots in Perennial Grass Experiments


First published on 14 August 2018

Pubmed ID: 30102263

Keywords: No keywords in Pubmed

McPherson MR, Wang P, Marsh EL, Mitchell RB, Schachtman DP

Plant and soil microbiome studies are becoming increasingly important for understanding the roles microorganisms play in agricultural productivity. The purpose of this manuscript is to provide detail on how to rapidly sample soil, rhizosphere, and endosphere of replicated field trials and analyze changes that may occur in the microbial communities due to sample type, treatment, and plant genotype. The experiment used to demonstrate these methods consists of replicated field plots containing two, pure, warm-season grasses (Panicum virgatum and Andropogon gerardii) and a low-diversity grass mixture (A. gerardii, Sorghastrum nutans, and Bouteloua curtipendula). Briefly, plants are excavated, a variety of roots are cut and placed in phosphate buffer, and then shaken to collect the rhizosphere. Roots are brought to the laboratory on ice and surface sterilized with bleach and ethanol (EtOH). The rhizosphere is filtered and concentrated by centrifugation. Excavated soil from around the root ball is placed into plastic bags and brought to the lab where a small amount of soil is taken for DNA extractions. DNA is extracted from roots, soil, and rhizosphere and then amplified with primers for the V4 region of the 16S rRNA gene. Amplicons are sequenced, then analyzed with open access bioinformatics tools. These methods allow researchers to test how the microbial community diversity and composition varies due to sample type, treatment, and plant genotype. Using these methods along with statistical models, the representative results demonstrate there are significant differences in microbial communities of roots, rhizosphere, and soil. Methods presented here provide a complete set of steps for how to collect field samples, isolate, extract, quantify, amplify, and sequence DNA, and analyze microbial community diversity and composition in replicated field trials.

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Long-Term Selection in Hybrid Sorghum Breeding Programs


First published on 12 August 2018

Pfeiffer BK, Pietsch D, Schnell RW, Rooney WL

Estimating genetic gains in sorghum [Sorghum bicolor (L.) Moench] is necessary to assess whether the current rates of improvement will meet future production demands. The current study was conducted to determine the rate of genetic gain in yield and associated traits over the commercial hybrid era using sorghum germplasm from the Texas A&M sorghum breeding program and a US sorghum seed company. From the Texas A&M AgriLife program, 60 hybrids that represented a 50-yr span of hybrid breeding were grown and evaluated in five environments across Texas in 2016. In a separate set of trials, 14 commercial hybrids representing a similar timespan were evaluated in three Texas environments in 2016. In both programs, grain sorghum yields increased 0.008 t ha−1 annually over their respective timespans. Traits that increased over time included yield potential per plant, heterosis, test weight, panicle size, and grain number per panicle, whereas leaf angle, days to maturity, plant height, and yield stability demonstrated little to no change. Overall, ∼60% of total yield gains in US sorghum production are attributed to genetic improvement through sorghum breeding. Compared with other major US field crops, the rate of genetic gain in sorghum has been slower, presumably due to a combination of factors, which include continually shifting production environments, changing priorities in traits, reduced research investments (compared with other crops), and less-than-optimized heterotic groups.

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A comparative transcriptional landscape of maize and sorghum obtained by single-molecule sequencing


First published on 2 May 2018

Pubmed ID: 29712755

Keywords: No keywords in Pubmed

Wang B, Regulski M, Tseng E, Olson A, Goodwin S, McCombie WR, Ware D

Maize and sorghum are both important crops with similar overall plant architectures, but they have key differences, especially in regard to their inflorescences. To better understand these two organisms at the molecular level, we compared expression profiles of both protein-coding and noncoding transcripts in 11 matched tissues using single-molecule, long-read, deep RNA sequencing. This comparative analysis revealed large numbers of novel isoforms in both species. Evolutionarily young genes were likely to be generated in reproductive tissues and usually had fewer isoforms than old genes. We also observed similarities and differences in alternative splicing patterns and activities, both among tissues and between species. The maize subgenomes exhibited no bias in isoform generation; however, genes in the B genome were more highly expressed in pollen tissue, whereas genes in the A genome were more highly expressed in endosperm. We also identified a number of splicing events conserved between maize and sorghum. In addition, we generated comprehensive and high-resolution maps of poly(A) sites, revealing similarities and differences in mRNA cleavage between the two species. Overall, our results reveal considerable splicing and expression diversity between sorghum and maize, well beyond what was reported in previous studies, likely reflecting the differences in architecture between these two species.

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Drought delays development of the sorghum root microbiome and enriches for monoderm bacteria


First published on 19 April 2018

Pubmed ID: 29666229

Keywords: drought, metatranscriptome, microbiome, root, sorghum

Xu L, Naylor D, Dong Z, Simmons T, Pierroz G, Hixson KK, Kim YM, Zink EM, Engbrecht KM, Wang Y, Gao C, DeGraaf S, Madera MA, Sievert JA, Hollingsworth J, Birdseye D, Scheller HV, Hutmacher R, Dahlberg J, Jansson C, Taylor JW, Lemaux PG, Coleman-Derr D

Drought stress is a major obstacle to crop productivity, and the severity and frequency of drought are expected to increase in the coming century. Certain root-associated bacteria have been shown to mitigate the negative effects of drought stress on plant growth, and manipulation of the crop microbiome is an emerging strategy for overcoming drought stress in agricultural systems, yet the effect of drought on the development of the root microbiome is poorly understood. Through 16S rRNA amplicon and metatranscriptome sequencing, as well as root metabolomics, we demonstrate that drought delays the development of the early sorghum root microbiome and causes increased abundance and activity of monoderm bacteria, which lack an outer cell membrane and contain thick cell walls. Our data suggest that altered plant metabolism and increased activity of bacterial ATP-binding cassette (ABC) transporter genes are correlated with these shifts in community composition. Finally, inoculation experiments with monoderm isolates indicate that increased colonization of the root during drought can positively impact plant growth. Collectively, these results demonstrate the role that drought plays in restructuring the root microbiome and highlight the importance of temporal sampling when studying plant-associated microbiomes.

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A cytochrome P450 CYP71 enzyme expressed in Sorghum bicolor root hair cells participates in the biosynthesis of the benzoquinone allelochemical sorgoleone


First published on 21 February 2018

Pubmed ID: 29461628

Keywords: Sorghum bicolor, CYP71AM1, allelochemical, cytochrome P450 monooxygenase, sorgoleone

Pan Z, Baerson SR, Wang M, Bajsa-Hirschel J, Rimando AM, Wang X, Nanayakkara NPD, Noonan BP, Fromm ME, Dayan FE, Khan IA, Duke SO

Sorgoleone, a major component of the hydrophobic root exudates of Sorghum spp., is probably responsible for many of the allelopathic properties attributed to members of this genus. Much of the biosynthetic pathway for this compound has been elucidated, with the exception of the enzyme responsible for the catalysis of the addition of two hydroxyl groups to the resorcinol ring. A library prepared from isolated Sorghum bicolor root hair cells was first mined for P450-like sequences, which were then analyzed by quantitative reverse transcription-polymerase chain reaction (RT-qPCR) to identify those preferentially expressed in root hairs. Full-length open reading frames for each candidate were generated, and then analyzed biochemically using both a yeast expression system and transient expression in Nicotiana benthamiana leaves. RNA interference (RNAi)-mediated repression in transgenic S. bicolor was used to confirm the roles of these candidates in the biosynthesis of sorgoleone in planta. A P450 enzyme, designated CYP71AM1, was found to be capable of catalyzing the formation of dihydrosorgoleone using 5-pentadecatrienyl resorcinol-3-methyl ether as substrate, as determined by gas chromatography-mass spectroscopy (GC-MS). RNAi-mediated repression of CYP71AM1 in S. bicolor resulted in decreased sorgoleone contents in multiple independent transformant events. Our results strongly suggest that CYP71AM1 participates in the biosynthetic pathway of the allelochemical sorgoleone.

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Whole-Genome Sequence Accuracy Is Improved by Replication in a Population of Mutagenized Sorghum


First published on 31 January 2018

Pubmed ID: 29378822

Keywords: EMS, SNP, accuracy, mutagenesis, mutants, mutations, polymorphisms, rare variants, sorghum

Addo-Quaye C, Tuinstra M, Carraro N, Weil C, Dilkes BP

The accurate detection of induced mutations is critical for both forward and reverse genetics studies. Experimental chemical mutagenesis induces relatively few single base changes per individual. In a complex eukaryotic genome, false positive detection of mutations can occur at or above this mutagenesis rate. We demonstrate here, using a population of ethyl methanesulfonate (EMS)-treated Sorghum bicolor BTx623 individuals, that using replication to detect false positive-induced variants in next-generation sequencing (NGS) data permits higher throughput variant detection with greater accuracy. We used a lower sequence coverage depth (average of 7×) from 586 independently mutagenized individuals and detected 5,399,493 homozygous single nucleotide polymorphisms (SNPs). Of these, 76% originated from only 57,872 genomic positions prone to false positive variant calling. These positions are characterized by high copy number paralogs where the error-prone SNP positions are at copies containing a variant at the SNP position. The ability of short stretches of homology to generate these error-prone positions suggests that incompletely assembled or poorly mapped repeated sequences are one driver of these error-prone positions. Removal of these false positives left 1,275,872 homozygous and 477,531 heterozygous EMS-induced SNPs, which, congruent with the mutagenic mechanism of EMS, were >98% G:C to A:T transitions. Through this analysis, we generated a collection of sequence indexed mutants of sorghum. This collection contains 4035 high-impact homozygous mutations in 3637 genes and 56,514 homozygous missense mutations in 23,227 genes. Each line contains, on average, 2177 annotated homozygous SNPs per genome, including seven likely gene knockouts and 96 missense mutations. The number of mutations in a transcript was linearly correlated with the transcript length and also the G+C count, but not with the GC/AT ratio. Analysis of the detected mutagenized positions identified CG-rich patches, and flanking sequences strongly influenced EMS-induced mutation rates. This method for detecting false positive-induced mutations is generally applicable to any organism, is independent of the choice of in silico variant-calling algorithm, and is most valuable when the true mutation rate is likely to be low, such as in laboratory-induced mutations or somatic mutation detection in medicine.

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The Sorghum bicolor reference genome: improved assembly, gene annotations, a transcriptome atlas, and signatures of genome organization


First published on 22 November 2017

Pubmed ID: 29161754

Keywords: Sorghum bicolor, discrete Fourier transform, gene annotation, genetic variation, genome assembly, kinase, nucleosome occupancy, reference genome, satellite DNA

McCormick RF, Truong SK, Sreedasyam A, Jenkins J, Shu S, Sims D, Kennedy M, Amirebrahimi M, Weers BD, McKinley B, Mattison A, Morishige DT, Grimwood J, Schmutz J, Mullet JE

Sorghum bicolor is a drought tolerant C4 grass used for the production of grain, forage, sugar, and lignocellulosic biomass and a genetic model for C4 grasses due to its relatively small genome (approximately 800 Mbp), diploid genetics, diverse germplasm, and colinearity with other C4 grass genomes. In this study, deep sequencing, genetic linkage analysis, and transcriptome data were used to produce and annotate a high-quality reference genome sequence. Reference genome sequence order was improved, 29.6 Mbp of additional sequence was incorporated, the number of genes annotated increased 24% to 34 211, average gene length and N50 increased, and error frequency was reduced 10-fold to 1 per 100 kbp. Subtelomeric repeats with characteristics of Tandem Repeats in Miniature (TRIM) elements were identified at the termini of most chromosomes. Nucleosome occupancy predictions identified nucleosomes positioned immediately downstream of transcription start sites and at different densities across chromosomes. Alignment of more than 50 resequenced genomes from diverse sorghum genotypes to the reference genome identified approximately 7.4 M single nucleotide polymorphisms (SNPs) and 1.9 M indels. Large-scale variant features in euchromatin were identified with periodicities of approximately 25 kbp. A transcriptome atlas of gene expression was constructed from 47 RNA-seq profiles of growing and developed tissues of the major plant organs (roots, leaves, stems, panicles, and seed) collected during the juvenile, vegetative and reproductive phases. Analysis of the transcriptome data indicated that tissue type and protein kinase expression had large influences on transcriptional profile clustering. The updated assembly, annotation, and transcriptome data represent a resource for C4 grass research and crop improvement.

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High‐throughput profiling and analysis of plant responses over time to abiotic stress


First published on 25 October 2017

Pubmed ID: 31245669

Keywords: abiotic stress, computational modeling, image analysis, ionomics, large‐scale biology, nitrogen stress

Veley KM, Berry JC, Fentress SJ, Schachtman DP, Baxter I, Bart R

Sorghum (Sorghum bicolor (L.) Moench) is a rapidly growing, high-biomass crop prized for abiotic stress tolerance. However, measuring genotype-by-environment (G x E) interactions remains a progress bottleneck. We subjected a panel of 30 genetically diverse sorghum genotypes to a spectrum of nitrogen deprivation and measured responses using high-throughput phenotyping technology followed by ionomic profiling. Responses were quantified using shape (16 measurable outputs), color (hue and intensity), and ionome (18 elements). We measured the speed at which specific genotypes respond to environmental conditions, in terms of both biomass and color changes, and identified individual genotypes that perform most favorably. With this analysis, we present a novel approach to quantifying color-based stress indicators over time. Additionally, ionomic profiling was conducted as an independent, low-cost, and high-throughput option for characterizing G x E, identifying the elements most affected by either genotype or treatment and suggesting signaling that occurs in response to the environment. This entire dataset and associated scripts are made available through an open-access, user-friendly, web-based interface. In summary, this work provides analysis tools for visualizing and quantifying plant abiotic stress responses over time. These methods can be deployed as a time-efficient method of dissecting the genetic mechanisms used by sorghum to respond to the environment to accelerate crop improvement.

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Dynamics of gene expression during development and expansion of vegetative stem internodes of bioenergy sorghum


First published on 27 June 2017

Pubmed ID: 28649278

Keywords: Bioenergy, Cell cycle, Hormone, Internode, Sorghum, Transcriptome

Kebrom TH, McKinley B, Mullet JE

BACKGROUND: Bioenergy sorghum accumulates 75% of shoot biomass in stem internodes. Grass stem internodes are formed during vegetative growth and elongate in response to developmental and environmental signals. To identify genes and molecular mechanisms that modulate the extent of internode growth, we conducted microscopic and transcriptomic analyses of four successive sub-apical vegetative internodes representing different stages of internode development of the bioenergy sorghum genotype R.07020.RESULTS: Stem internodes of sorghum genotype R.07020 are formed during the vegetative phase and their length is enhanced by environmental signals such as shade and floral induction in short days. During vegetative growth, the first visible and youngest sub-apical internode was ~0.7 cm in length, whereas the fourth fully expanded internode was ~5 cm in length. Microscopic analyses revealed that all internode tissue types including pith parenchyma and vascular bundles are present in the four successive internodes. Growth in the first two sub-apical internodes occurred primarily through an increase in cell number consistent with expression of genes involved in the cell cycle and DNA replication. Growth of the 3rd internode was associated with an increase in cell length and growth cessation in the 4th internode was associated with up-regulation of genes involved in secondary cell wall deposition. The expression of genes involved in hormone metabolism and signaling indicates that GA, BR, and CK activity decreased while ethylene, ABA, and JA increased in the 3rd/4th internodes. While the level of auxin appears to be increasing as indicated by the up-regulation of ARFs, down-regulation of TIR during development indicates that auxin signaling is also modified. The expression patterns of transcription factors are closely associated with their role during the development of the vegetative internodes.CONCLUSIONS: Microscopic and transcriptome analyses of four successive sub-apical internodes characterized the developmental progression of vegetative stem internodes from initiation through full elongation in the sorghum genotype R.07020. Transcriptome profiling indicates that dynamic variation in the levels and action of GA, CK, IAA, BR, ethylene, ABA, and JA modulate gene expression and growth during internode growth and development. This study provides detailed microscopic and transcriptomic data useful for identifying genes and molecular pathways regulating internode elongation in response to various developmental and environmental signals.

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DNA methylation and gene expression regulation associated with vascularization in Sorghum bicolor


First published on 12 February 2017

Pubmed ID: 28186631

Keywords: biofuel, cell type-specific, epigenetics, sorghum (Sorghum bicolor), transcriptome

Turco GM, Kajala K, Kunde-Ramamoorthy G, Ngan CY, Olson A, Deshphande S, Tolkunov D, Waring B, Stelpflug S, Klein P, Schmutz J, Kaeppler S, Ware D, Wei CL, Etchells JP, Brady SM

Plant secondary cell walls constitute the majority of plant biomass. They are predominantly found in xylem cells, which are derived from vascular initials during vascularization. Little is known about these processes in grass species despite their emerging importance as biomass feedstocks. The targeted biofuel crop Sorghum bicolor has a sequenced and well-annotated genome, making it an ideal monocot model for addressing vascularization and biomass deposition. Here we generated tissue-specific transcriptome and DNA methylome data from sorghum shoots, roots and developing root vascular and nonvascular tissues. Many genes associated with vascular development in other species show enriched expression in developing vasculature. However, several transcription factor families varied in vascular expression in sorghum compared with Arabidopsis and maize. Furthermore, differential expression of genes associated with DNA methylation were identified between vascular and nonvascular tissues, implying that changes in DNA methylation are a feature of sorghum root vascularization, which we confirmed using tissue-specific DNA methylome data. Roots treated with a DNA methylation inhibitor also showed a significant decrease in root length. Tissues and organs can be discriminated based on their genomic methylation patterns and methylation context. Consequently, tissue-specific changes in DNA methylation are part of the normal developmental process.

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Forward Genetics by Sequencing EMS Variation-Induced Inbred Lines


First published on 4 January 2017

Pubmed ID: 28040779

Keywords: SNP, Sorghum, gibberellin, mapping, mutant

Addo-Quaye C, Buescher E, Best N, Chaikam V, Baxter I, Dilkes BP

In order to leverage novel sequencing techniques for cloning genes in eukaryotic organisms with complex genomes, the false positive rate of variant discovery must be controlled for by experimental design and informatics. We sequenced five lines from three pedigrees of ethyl methanesulfonate (EMS)-mutagenized Sorghum bicolor, including a pedigree segregating a recessive dwarf mutant. Comparing the sequences of the lines, we were able to identify and eliminate error-prone positions. One genomic region contained EMS mutant alleles in dwarfs that were homozygous reference sequences in wild-type siblings and heterozygous in segregating families. This region contained a single nonsynonymous change that cosegregated with dwarfism in a validation population and caused a premature stop codon in the Sorghum ortholog encoding the gibberellic acid (GA) biosynthetic enzyme ent-kaurene oxidase. Application of exogenous GA rescued the mutant phenotype. Our method for mapping did not require outcrossing and introduced no segregation variance. This enables work when line crossing is complicated by life history, permitting gene discovery outside of genetic models. This inverts the historical approach of first using recombination to define a locus and then sequencing genes. Our formally identical approach first sequences all the genes and then seeks cosegregation with the trait. Mutagenized lines lacking obvious phenotypic alterations are available for an extension of this approach: mapping with a known marker set in a line that is phenotypically identical to starting material for EMS mutant generation.

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A Genomic Resource for the Development, Improvement, and Exploitation of Sorghum for Bioenergy


First published on 1 July 2016

Pubmed ID: 27356613

Keywords: Bioenergy Association Panel, MPP, Multiparent Advanced Generation Inter-Cross (MAGIC), biomass composition, carbon partitioning, multiparental populations, nonstructural sugars

Brenton ZW, Cooper EA, Myers MT, Boyles RE, Shakoor N, Zielinski KJ, Rauh BL, Bridges WC, Morris GP, Kresovich S

With high productivity and stress tolerance, numerous grass genera of the Andropogoneae have emerged as candidates for bioenergy production. To optimize these candidates, research examining the genetic architecture of yield, carbon partitioning, and composition is required to advance breeding objectives. Significant progress has been made developing genetic and genomic resources for Andropogoneae, and advances in comparative and computational genomics have enabled research examining the genetic basis of photosynthesis, carbon partitioning, composition, and sink strength. To provide a pivotal resource aimed at developing a comparative understanding of key bioenergy traits in the Andropogoneae, we have established and characterized an association panel of 390 racially, geographically, and phenotypically diverse Sorghum bicolor accessions with 232,303 genetic markers. Sorghum bicolor was selected because of its genomic simplicity, phenotypic diversity, significant genomic tools, and its agricultural productivity and resilience. We have demonstrated the value of sorghum as a functional model for candidate gene discovery for bioenergy Andropogoneae by performing genome-wide association analysis for two contrasting phenotypes representing key components of structural and non-structural carbohydrates. We identified potential genes, including a cellulase enzyme and a vacuolar transporter, associated with increased non-structural carbohydrates that could lead to bioenergy sorghum improvement. Although our analysis identified genes with potentially clear functions, other candidates did not have assigned functions, suggesting novel molecular mechanisms for carbon partitioning traits. These results, combined with our characterization of phenotypic and genetic diversity and the public accessibility of each accession and genomic data, demonstrate the value of this resource and provide a foundation for future improvement of sorghum and related grasses for bioenergy production.

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The Ensembl gene annotation system


First published on 25 June 2016

Pubmed ID: 27337980

Keywords: No keywords in Pubmed

Aken BL, Ayling S, Barrell D, Clarke L, Curwen V, Fairley S, Fernandez Banet J, Billis K, García Girón C, Hourlier T, Howe K, Kähäri A, Kokocinski F, Martin FJ, Murphy DN, Nag R, Ruffier M, Schuster M, Tang YA, Vogel JH, White S, Zadissa A, Flicek P, Searle SM

The Ensembl gene annotation system has been used to annotate over 70 different vertebrate species across a wide range of genome projects. Furthermore, it generates the automatic alignment-based annotation for the human and mouse GENCODE gene sets. The system is based on the alignment of biological sequences, including cDNAs, proteins and RNA-seq reads, to the target genome in order to construct candidate transcript models. Careful assessment and filtering of these candidate transcripts ultimately leads to the final gene set, which is made available on the Ensembl website. Here, we describe the annotation process in detail.Database URL: http://www.ensembl.org/index.html.

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Independent and Parallel Evolution of New Genes by Gene Duplication in Two Origins of C4 Photosynthesis Provides New Insight into the Mechanism of Phloem Loading in C4 Species


First published on 27 March 2016

Pubmed ID: 27016024

Keywords: No keywords in Pubmed

Emms DM, Covshoff S, Hibberd JM, Kelly S

UNLABELLED: C4 photosynthesis is considered one of the most remarkable examples of evolutionary convergence in eukaryotes. However, it is unknown whether the evolution of C4 photosynthesis required the evolution of new genes. Genome-wide gene-tree species-tree reconciliation of seven monocot species that span two origins of C4 photosynthesis revealed that there was significant parallelism in the duplication and retention of genes coincident with the evolution of C4 photosynthesis in these lineages. Specifically, 21 orthologous genes were duplicated and retained independently in parallel at both C4 origins. Analysis of this gene cohort revealed that the set of parallel duplicated and retained genes is enriched for genes that are preferentially expressed in bundle sheath cells, the cell type in which photosynthesis was activated during C4 evolution. Furthermore, functional analysis of the cohort of parallel duplicated genes identified SWEET-13 as a potential key transporter in the evolution of C4 photosynthesis in grasses, and provides new insight into the mechanism of phloem loading in these C4 species.KEY WORDS: C4 photosynthesis, gene duplication, gene families, parallel evolution.

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Natural Variation in Synthesis and Catabolism Genes Influences Dhurrin Content in Sorghum


First published on 1 July 2015

Chad M. Hayes Gloria B. Burow Patrick J. Brown Carrie Thurber Zhanguo Xin John J. Burke

Cyanogenic glucosides are natural compounds found in more than 1000 species of angiosperms that produce HCN and are deemed undesirable for agricultural use. However, these compounds are important components of the primary defensive mechanisms of many plant species. One of the best‐studied cyanogenic glucosides is dhurrin [(S)‐p‐hydroxymandelonitrile‐β‐D‐glucopyranoside], which is produced primarily in sorghum [Sorghum bicolor (L.) Moench]. The biochemical basis for dhurrin metabolism is well established; however, little information is available on its genetic control. Here, we dissect the genetic control of leaf dhurrin content through a genome‐wide association study (GWAS) using a panel of 700 diverse converted sorghum lines (conversion panel) previously subjected to pre‐breeding and selected for short stature (∼1 m in height) and photoperiod insensitivity. The conversion panel was grown for 2 yr in three environments. Wide variation for leaf dhurrin content was found in the sorghum conversion panel, with the Caudatum group exhibiting the highest dhurrin content and the Guinea group showing the lowest dhurrin content. A GWAS using a mixed linear model revealed significant associations (a false discovery rate [FDR] < 0.05) close to both UGT 185B1 in the canonical biosynthetic gene cluster on chromosome 1 and close to the catabolic dhurrinase loci on chromosome 8. Dhurrin content was associated consistently with biosynthetic genes in the two N‐fertilized environments, while dhurrin content was associated with catabolic loci in the environment without supplemental N. These results suggest that genes for both biosynthesis and catabolism are important in determining natural variation for leaf dhurrin in sorghum in different environments.

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MOROKOSHI: transcriptome database in Sorghum bicolor


First published on 17 December 2014

Pubmed ID: 25505007

Keywords: Database, FL-cDNA, NGS, New transcript, Plant, Sorghum

Makita Y, Shimada S, Kawashima M, Kondou-Kuriyama T, Toyoda T, Matsui M

In transcriptome analysis, accurate annotation of each transcriptional unit and its expression profile is essential. A full-length cDNA (FL-cDNA) collection facilitates the refinement of transcriptional annotation, and accurate transcription start sites help to unravel transcriptional regulation. We constructed a normalized FL-cDNA library from eight growth stages of aerial tissues in Sorghum bicolor and isolated 37,607 clones. These clones were Sanger sequenced from the 5' and/or 3' ends and in total 38,981 high-quality expressed sequence tags (ESTs) were obtained. About one-third of the transcripts of known genes were captured as FL-cDNA clone resources. In addition to these, we also annotated 272 novel genes, 323 antisense transcripts and 1,672 candidate isoforms. These clones are available from the RIKEN Bioresource Center. After obtaining accurate annotation of transcriptional units, we performed expression profile analysis. We carried out spikelet-, seed- and stem-specific RNA sequencing (RNA-Seq) analysis and confirmed the expression of 70.6% of the newly identified genes. We also downloaded 23 sorghum RNA-Seq samples that are publicly available and these are shown on a genome browser together with our original FL-cDNA and RNA-Seq data. Using our original and publicly available data, we made an expression profile of each gene and identified the top 20 genes with the most similar expression. In addition, we visualized their relationships in gene co-expression networks. Users can access and compare various transcriptome data from S, bicolor at http://sorghum.riken.jp.

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Identification of differentially expressed genes between sorghum genotypes with contrasting nitrogen stress tolerance by genome-wide transcriptional profiling


First published on 7 March 2014

Pubmed ID: 24597475

Keywords: No keywords in Pubmed

Gelli M, Duo Y, Konda AR, Zhang C, Holding D, Dweikat I

BACKGROUND: Sorghum is an important cereal crop, which requires large quantities of nitrogen fertilizer for achieving commercial yields. Identification of the genes responsible for low-N tolerance in sorghum will facilitate understanding of the molecular mechanisms of low-N tolerance, and also facilitate the genetic improvement of sorghum through marker-assisted selection or gene transformation. In this study we compared the transcriptomes of root tissues from seven sorghum genotypes having differential response to low-N stress.RESULTS: Illumina RNA-sequencing detected several common differentially expressed genes (DEGs) between four low-N tolerant sorghum genotypes (San Chi San, China17, KS78 and high-NUE bulk) and three sensitive genotypes (CK60, BTx623 and low-NUE bulk). In sensitive genotypes, N-stress increased the abundance of DEG transcripts associated with stress responses including oxidative stress and stimuli were abundant. The tolerant genotypes adapt to N deficiency by producing greater root mass for efficient uptake of nutrients. In tolerant genotypes, higher abundance of transcripts related to high affinity nitrate transporters (NRT2.2, NRT2.3, NRT2.5, and NRT2.6) and lysine histidine transporter 1 (LHT1), may suggest an improved uptake efficiency of inorganic and organic forms of nitrogen. Higher abundance of SEC14 cytosolic factor family protein transcript in tolerant genotypes could lead to increased membrane stability and tolerance to N-stress.CONCLUSIONS: Comparison of transcriptomes between N-stress tolerant and sensitive genotypes revealed several common DEG transcripts. Some of these DEGs were evaluated further by comparing the transcriptomes of genotypes grown under full N. The DEG transcripts showed higher expression in tolerant genotypes could be used for transgenic over-expression in sensitive genotypes of sorghum and related crops for increased tolerance to N-stress, which results in increased nitrogen use efficiency for sustainable agriculture.

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Dissecting Genome-Wide Association Signals for Loss-Of-Function Phenotypes in Sorghum Flavonoid Pigmentation Traits


First published on 21 September 2013

Pubmed ID: 24048646

Keywords: genome scan, grain pigmentation, null alleles, quantitative trait loci, structured populations

Morris GP, Rhodes DH, Brenton Z, Ramu P, Thayil VM, Deshpande S, Hash CT, Acharya C, Mitchell SE, Buckler ES, Yu J, Kresovich S

Genome-wide association studies are a powerful method to dissect the genetic basis of traits, although in practice the effects of complex genetic architecture and population structure remain poorly understood. To compare mapping strategies we dissected the genetic control of flavonoid pigmentation traits in the cereal grass sorghum by using high-resolution genotyping-by-sequencing single-nucleotide polymorphism markers. Studying the grain tannin trait, we find that general linear models (GLMs) are not able to precisely map tan1-a, a known loss-of-function allele of the Tannin1 gene, with either a small panel (n = 142) or large association panel (n = 336), and that indirect associations limit the mapping of the Tannin1 locus to Mb-resolution. A GLM that accounts for population structure (Q) or standard mixed linear model that accounts for kinship (K) can identify tan1-a, whereas a compressed mixed linear model performs worse than the naive GLM. Interestingly, a simple loss-of-function genome scan, for genotype-phenotype covariation only in the putative loss-of-function allele, is able to precisely identify the Tannin1 gene without considering relatedness. We also find that the tan1-a allele can be mapped with gene resolution in a biparental recombinant inbred line family (n = 263) using genotyping-by-sequencing markers but lower precision in the mapping of vegetative pigmentation traits suggest that consistent gene-level resolution will likely require larger families or multiple recombinant inbred lines. These findings highlight that complex association signals can emerge from even the simplest traits given epistasis and structured alleles, but that gene-resolution mapping of these traits is possible with high marker density and appropriate models.

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Retrospective genomic analysis of sorghum adaptation to temperate-zone grain production


First published on 28 June 2013

Pubmed ID: 23803286

Keywords: No keywords in Pubmed

Thurber CS, Ma JM, Higgins RH, Brown PJ

BACKGROUND: Sorghum is a tropical C4 cereal that recently adapted to temperate latitudes and mechanized grain harvest through selection for dwarfism and photoperiod-insensitivity. Quantitative trait loci for these traits have been introgressed from a dwarf temperate donor into hundreds of diverse sorghum landraces to yield the Sorghum Conversion lines. Here, we report the first comprehensive genomic analysis of the molecular changes underlying this adaptation.RESULTS: We apply genotyping-by-sequencing to 1,160 Sorghum Conversion lines and their exotic progenitors, and map donor introgressions in each Sorghum Conversion line. Many Sorghum Conversion lines carry unexpected haplotypes not found in either presumed parent. Genome-wide mapping of introgression frequencies reveals three genomic regions necessary for temperate adaptation across all Sorghum Conversion lines, containing the Dw1, Dw2, and Dw3 loci on chromosomes 9, 6, and 7 respectively. Association mapping of plant height and flowering time in Sorghum Conversion lines detects significant associations in the Dw1 but not the Dw2 or Dw3 regions. Subpopulation-specific introgression mapping suggests that chromosome 6 contains at least four loci required for temperate adaptation in different sorghum genetic backgrounds. The Dw1 region fractionates into separate quantitative trait loci for plant height and flowering time.CONCLUSIONS: Generating Sorghum Conversion lines has been accompanied by substantial unintended gene flow. Sorghum adaptation to temperate-zone grain production involves a small number of genomic regions, each containing multiple linked loci for plant height and flowering time. Further characterization of these loci will accelerate the adaptation of sorghum and related grasses to new production systems for food and fuel.

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Comparative transcriptomics of three Poaceae species reveals patterns of gene expression evolution


First published on 1 August 2012

Pubmed ID: 22443345

Keywords: Sorghum, transcriptomics, rice, Poaceae, gene expression

Rebecca M Davidson, Malali Gowda, Gaurav Moghe, Haining Lin, Brieanne Vaillancourt, Shin-Han Shiu, Ning Jiang, C Robin Buell

The Poaceae family, also known as the grasses, includes agronomically important cereal crops such as rice, maize, sorghum, and wheat. Previous comparative studies have shown that much of the gene content is shared among the grasses; however, functional conservation of orthologous genes has yet to be explored. To gain an understanding of the genome-wide patterns of evolution of gene expression across reproductive tissues, we employed a sequence-based approach to compare analogous transcriptomes in species representing three Poaceae subgroups including the Pooideae (Brachypodium distachyon), the Panicoideae (sorghum), and the Ehrhartoideae (rice). Our transcriptome analyses reveal that only a fraction of orthologous genes exhibit conserved expression patterns. A high proportion of conserved orthologs include genes that are upregulated in physiologically similar tissues such as leaves, anther, pistil, and embryo, while orthologs that are highly expressed in seeds show the most diverged expression patterns. More generally, we show that evolution of gene expression profiles and coding sequences in the grasses may be linked. Genes that are highly and broadly expressed tend to be conserved at the coding sequence level while genes with narrow expression patterns show accelerated rates of sequence evolution. We further show that orthologs in syntenic genomic blocks are more likely to share correlated expression patterns compared with non-syntenic orthologs. These findings are important for agricultural improvement because sequence information is transferred from model species, such as Brachypodium, rice, and sorghum to crop plants without sequenced genomes.

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Phytozome: a comparative platform for green plant genomics


First published on 24 November 2011

Pubmed ID: 22110026

Keywords: No keywords in Pubmed

Goodstein DM, Shu S, Howson R, Neupane R, Hayes RD, Fazo J, Mitros T, Dirks W, Hellsten U, Putnam N, Rokhsar DS

The number of sequenced plant genomes and associated genomic resources is growing rapidly with the advent of both an increased focus on plant genomics from funding agencies, and the application of inexpensive next generation sequencing. To interact with this increasing body of data, we have developed Phytozome (http://www.phytozome.net), a comparative hub for plant genome and gene family data and analysis. Phytozome provides a view of the evolutionary history of every plant gene at the level of sequence, gene structure, gene family and genome organization, while at the same time providing access to the sequences and functional annotations of a growing number (currently 25) of complete plant genomes, including all the land plants and selected algae sequenced at the Joint Genome Institute, as well as selected species sequenced elsewhere. Through a comprehensive plant genome database and web portal, these data and analyses are available to the broader plant science research community, providing powerful comparative genomics tools that help to link model systems with other plants of economic and ecological importance.

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Genome-wide patterns of genetic variation in sweet and grain sorghum (Sorghum bicolor)


First published on 23 November 2011

Pubmed ID: 22104744

Keywords: No keywords in Pubmed

Zheng LY, Guo XS, He B, Sun LJ, Peng Y, Dong SS, Liu TF, Jiang S, Ramachandran S, Liu CM, Jing HC

BACKGROUND: Sorghum (Sorghum bicolor) is globally produced as a source of food, feed, fiber and fuel. Grain and sweet sorghums differ in a number of important traits, including stem sugar and juice accumulation, plant height as well as grain and biomass production. The first whole genome sequence of a grain sorghum is available, but additional genome sequences are required to study genome-wide and intraspecific variation for dissecting the genetic basis of these important traits and for tailor-designed breeding of this important C4 crop.RESULTS: We resequenced two sweet and one grain sorghum inbred lines, and identified a set of nearly 1,500 genes differentiating sweet and grain sorghum. These genes fall into ten major metabolic pathways involved in sugar and starch metabolisms, lignin and coumarin biosynthesis, nucleic acid metabolism, stress responses and DNA damage repair. In addition, we uncovered 1,057,018 SNPs, 99,948 indels of 1 to 10 bp in length and 16,487 presence/absence variations as well as 17,111 copy number variations. The majority of the large-effect SNPs, indels and presence/absence variations resided in the genes containing leucine rich repeats, PPR repeats and disease resistance R genes possessing diverse biological functions or under diversifying selection, but were absent in genes that are essential for life.CONCLUSIONS: This is a first report of the identification of genome-wide patterns of genetic variation in sorghum. High-density SNP and indel markers reported here will be a valuable resource for future gene-phenotype studies and the molecular breeding of this important crop and related species.

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


First published on 20 October 2011

Pubmed ID: 22008187

Keywords: No keywords in Pubmed

Dugas DV, Monaco MK, Olsen A, Klein RR, Kumari S, Ware D, Klein PE

BACKGROUND: Higher 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 sorghum's tolerance to water-limiting environments with a long-term aim of developing strategies to improve plant productivity under drought.RESULTS: RNA-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.CONCLUSIONS: The 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.

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Developing a Mini Core Collection of Sorghum for Diversified Utilization of Germplasm


First published on 1 September 2009

H. D. Upadhyaya R. P. S. Pundir S. L. Dwivedi C. L.L. Gowda V. G. Reddy S. Singh

The sorghum [Sorghum bicolor (L.) Moench] germplasm collection at the ICRISAT gene bank exceeds 37,000 accessions. A core collection of 2247 accessions was developed in 2001 to enable researchers to have access to a smaller set of germplasm. However, this core collection was found to be too large. To overcome this, a sorghum mini core (10% accessions of the core or 1% of the entire collection) was developed from the existing core collection. The core collection was evaluated for 11 qualitative and 10 quantitative traits in an augmented design using three control cultivars in the 2004–2005 post‐rainy season. The hierarchical cluster analysis of data using phenotypic distances resulted in 21 clusters. From each cluster, about 10% or a minimum of one accession was selected to form a mini core that comprised 242 accessions. The data in the mini core and core collections were compared using statistical parameters such as homogeneity of distribution for geographical origin, biological races, qualitative traits, means, variances, phenotypic diversity indices, and phenotypic correlations. These tests revealed that the mini core collection represented the core collection, which can be evaluated extensively for agronomic traits including resistance to biotic and abiotic stresses to identify accessions with desirable characteristics for use in crop improvement research and genomic studies.

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Applying genotyping (TILLING) and phenotyping analyses to elucidate gene function in a chemically induced sorghum mutant population


First published on 16 October 2008

Pubmed ID: 18854043

Keywords: No keywords in Pubmed

Xin Z, Wang ML, Barkley NA, Burow G, Franks C, Pederson G, Burke J

BACKGROUND: Sorghum [Sorghum bicolor (L.) Moench] is ranked as the fifth most important grain crop and serves as a major food staple and fodder resource for much of the world, especially in arid and semi-arid regions. The recent surge in sorghum research is driven by its tolerance to drought/heat stresses and its strong potential as a bioenergy feedstock. Completion of the sorghum genome sequence has opened new avenues for sorghum functional genomics. However, the availability of genetic resources, specifically mutant lines, is limited. Chemical mutagenesis of sorghum germplasm, followed by screening for mutants altered in important agronomic traits, represents a rapid and effective means of addressing this limitation. Induced mutations in novel genes of interest can be efficiently assessed using the technique known as Targeting Induced Local Lesion IN Genomes (TILLING).RESULTS: A sorghum mutant population consisting of 1,600 lines was generated from the inbred line BTx623 by treatment with the chemical agent ethyl methanesulfonate (EMS). Numerous phenotypes with altered morphological and agronomic traits were observed from M2 and M3 lines in the field. A subset of 768 mutant lines was analyzed by TILLING using four target genes. A total of five mutations were identified resulting in a calculated mutation density of 1/526 kb. Two of the mutations identified by TILLING and verified by sequencing were detected in the gene encoding caffeic acid O-methyltransferase (COMT) in two independent mutant lines. The two mutant lines segregated for the expected brown midrib (bmr) phenotype, a trait associated with altered lignin content and increased digestibility.CONCLUSION: TILLING as a reverse genetic approach has been successfully applied to sorghum. The diversity of the mutant phenotypes observed in the field, and the density of induced mutations calculated from TILLING indicate that this mutant population represents a useful resource for members of the sorghum research community. Moreover, TILLING has been demonstrated to be applicable for sorghum functional genomics by evaluating a small subset of the EMS-induced mutant lines.

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Community Resources and Strategies for Association Mapping in Sorghum


First published on 1 January 2008

Alexandra M. Casa Gael Pressoir Patrick J. Brown Sharon E. Mitchell William L. Rooney Mitchell R. Tuinstra Cleve D. Franks Stephen Kresovich

Association mapping is a powerful strategy for identifying genes underlying quantitative traits in plants. We have assembled and characterized genetic and phenotypic diversity of a sorghum [Sorghum bicolor (L.) Moench] panel suitable for association mapping, comprised of 377 accessions representing all major cultivated races (tropical lines from diverse geographic and climatic regions), and important U.S. breeding lines and their progenitors. Accessions were phenotyped for eight traits, and levels of population structure and familial relatedness were assessed with 47 simple sequence repeat (SSR) loci. The panel exhibited substantial morphological variation and little genotypic differentiation was observed between the converted tropical and breeding lines. The phenotypic and genotypic data were used to evaluate the performance of several association models in controlling for spurious associations. Our analysis indicated that association models that accounted for both population structure and kinship performed better than those that did not. In addition, we found that the optimal number of subpopulations used to correct for population structure was trait dependent. Although augmentation of the genotypic data with additional SSR loci may be necessary, the association models, genotypic data, and germplasm panel described here provide a starting point for sorghum researchers to begin association studies of traits and markers or candidate genes of interest.

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Prospection de Mils pénicillaires et Sorghos en Afrique de l’Ouest : Campagne 1976 : Niger


First published on 1 January 1976

Borgel Alain, Séquier Jacques

Searching for pearl millet and sorghum in West Africa: 1976 Campain: Niger

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Expanding and Vetting Sorghum bicolor Gene Annotations through Transcriptome and Methylome Sequencing


First published on 1 January 1970

Andrew Olson, Robert R. Klein, Diana V. Dugas, Zhenyuan Lu, Michael Regulski, Patricia E. Klein, Doreen Ware

With the emergence and subsequent advancement of next-generation sequence technology, detailed structural and functional characterization of genomes is readily attainable. Here, we have sampled the Sorghum bicolor methylome by shallow sequencing of HSO3– (bisulfite)-treated DNA and have used these data to identify methylation patterns associated with high confidence gene models. We trained a classifier to predict functional gene models based on expression levels, methylation profiles, and sequence conservation. We have expanded the transcriptome atlas by sequencing RNA from meristematic tissues, florets, and embryos, and utilized this information to develop a more complete annotation of the sorghum transcriptome. Our gene annotations modify 60% of Sbi1.4 (version 1.4 of sorghum gene annotations) gene models. The updated models most often have extended untranslated region (UTR) annotations (18,105), but some show longer protein coding regions (5096) or previously unannotated alternative transcripts (6493). A phylogenetic analysis suggests that 800 genes are missing from annotation Sbi1.4 and 400 gene models are split. The new annotations resolve 50% of split gene models and include 30% of conserved genes missing from the Sbi1.4 annotation. Using our classifier, we identified a large set of 34,276 novel potentially functional transcribed regions. These transcribed regions include protein coding genes, non-coding RNAs, and other classes of gene products.

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Core Collection of Sorghum : II. Comparison of Three Random Sampling Strategies


First published on 1 January 1970

C. Grenier P. Hamon P.J. Bramel‐Cox

Since 1972, the International Crops Research Institute for the Semi‐Arid Tropics (ICRISAT) has maintained a large collection of sorghum in India. The collection size has continuously increased, and the total number of accessions at present conserved in the gene bank has reached about 36 000 accessions. The need to help management was considered, and this study was conducted to establish core collections. This sorghum collection was earlier stratified into four clusters according to the photoperiod sensitivity. Then, considering the core collection strategy, we used three random sampling procedures to determine the specific accessions to be included in the core [i.e., a constant portion (Core C), a proportional (Core P), and a proportional to the logarithm (Core L)] of the photoperiod group size sampling strategy. Both the Core C and L were significantly different from the landrace collection with better representation of the smallest groups, such as landraces insensitive to photoperiod. Despite differences between the three core collections, estimates of global diversity through the Shannon‐Weaver Diversity Indices were of the same magnitude as the landrace collection. When compared, the Core C and L were significantly different. Core L sampled better for the characters, the race, and the latitudinal classes that were related to the photoperiod‐sensitive landraces. Thus, for establishing a core collection with the widest range of adaptation to photoperiod, we propose the use of a logarithmic sampling strategy, which identifies a broadly adapted set of genotypes.

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