Optimal flowering time is critical to the success of modern agriculture. Sorghum is a short-day tropical species that exhibits substantial photoperiod sensitivity and delayed flowering in long days. Genotypes with reduced photoperiod sensitivity enabled sorghum's utilization as a grain crop in temperate zones worldwide. In the present study, Ma 1 , the major repressor of sorghum flowering in long days, was identified as the pseudoresponse regulator protein 37 (PRR37) through positional cloning and analysis of SbPRR37 alleles that modulate flowering time in grain and energy sorghum. Several allelic variants of SbPRR37 were identified in early flowering grain sorghum germplasm that contain unique loss-of-function mutations. We show that in long days SbPRR37 activates expression of the floral inhibitor CONSTANS and represses expression of the floral activators Early Heading Date 1, FLOWERING LOCUS T, Zea mays CENTRORADIALIS 8, and floral induction. Expression of SbPRR37 is light dependent and regulated by the circadian clock, with peaks of RNA abundance in the morning and evening in long days. In short days, the evening-phase expression of SbPRR37 does not occur due to darkness, allowing sorghum to flower in this photoperiod. This study provides insight into an external coincidence mechanism of photoperiodic regulation of flowering time mediated by PRR37 in the short-day grass sorghum and identifies important alleles of SbPRR37 that are critical for the utilization of this tropical grass in temperate zone grain and bioenergy production.is a C4 grass native to Africa that provides an indispensable food source for over 300 million people inhabiting food-insecure regions worldwide (1). Although primarily grown for its grain and forage, highbiomass sorghum is also an excellent drought-tolerant energy crop for sustainable production of lignocellulosic-based biofuels (2). Forage and energy sorghums are selected for delayed flowering to increase biomass yield through longer duration of vegetative growth, whereas grain sorghums are selected for early flowering to ensure sufficient time for grain maturation and to avoid drought and frost. Optimal production of each of these sorghum crops requires the precise regulation of flowering time, which varies depending on planting location and climate. Differences in photoperiod sensitivity confer a wide range of flowering times on diverse accessions of the sorghum germplasm collection (3). Due to its critical importance to crop yield and hybrid seed production, photoperiodic regulation of flowering has been an important trait characterized by sorghum improvement programs dating back to the early 1900s (4).In Arabidopsis, flowering is induced in long days (LD) that expose plants to light in the evening during a phase of circadian clock oscillation required for induction of floral genes, consistent with the external coincidence model (5-7). Rhythmic expression of the core circadian clock components CIRCADIAN CLOCK ASSOCIATED 1 (CCA1)/LATE ELONGATED HYPOCOTYL (LHY) and TIMING OF CAB 1...

Sorghum is an important source of food, feed, and biofuel, especially in the semi-arid tropics because this cereal is well adapted to harsh, drought-prone environments. Post-flowering drought adaptation in sorghum is associated with the stay-green phenotype. Alleles that contribute to this complex trait have been mapped to four major QTL, Stg1-Stg4, using a population derived from BTx642 and RTx7000. Near-isogenic RTx7000 lines containing BTx642 DNA spanning one or more of the four stay-green QTL were constructed. The size and location of BTx642 DNA regions in each RTx7000 NIL were analysed using 62 DNA markers spanning the four stay-green QTL. RTx7000 NILs were identified that contained BTx642 DNA completely or partially spanning Stg1, Stg2, Stg3, or Stg4. NILs were also identified that contained sub-portions of each QTL and various combinations of the four major stay-green QTL. Physiological analysis of four RTx7000 NILs containing only Stg1, Stg2, Stg3, or Stg4 showed that BTx642 alleles in each of these loci could contribute to the stay-green phenotype. RTx7000 NILs containing BTx642 DNA corresponding to Stg2 retained more green leaf area at maturity under terminal drought conditions than RTx7000 or the other RTx7000 NILs. Under post-anthesis water deficit, a trend for delayed onset of leaf senescence compared with RTx7000 was also exhibited by the Stg2, Stg3, and Stg4 NILs, while significantly lower rates of leaf senescence in relation to RTx7000 were displayed by all of the Stg NILs to varying degrees, but particularly by the Stg2 NIL. Greener leaves at anthesis relative to RTx7000, indicated by higher SPAD values, were exhibited by the Stg1 and Stg4 NILs. The RTx7000 NILs created in this study provide the starting point for in-depth analysis of stay-green physiology, interaction among stay-green QTL and map-based cloning of the genes that underlie this trait.

Transcription in lysed barley plastids and Northern slot blot analyses were used to determine the relationship between changes in RNA levels and transcription during plastid development. Transcription in plastids of 4.5-9-day-old dark-grown or illuminated barley seedlings declined up to 10-fold as a function of plant age. Decreased transcription of some plastid genes (rbcL, psaA-psaB) was paralleled by decreased levels of mRNA. In other cases (16SrDNA, psbA) the changes in transcription were not followed by proportional changes in RNA levels indicating that RNA stability is important in establishing the amount of plastid RNA for these genes. Further analysis showed that transcription of the plastid rRNA transcription unit is regulated differently than the transcription of protein coding genes such as psbA or rbcL.

Sorghum is an important target for plant genomic mapping because of its adaptation to harsh environments, diverse germplasm collection, and value for comparing the genomes of grass species such as corn and rice. The construction of an integrated genetic and physical map of the sorghum genome (750 Mbp) is a primary goal of our sorghum genome project. To help accomplish this task, we have developed a new high-throughput PCR-based method for building BAC contigs and locating BAC clones on the sorghum genetic map. This task involved pooling 24,576 sorghum BAC clones (∼4× genome equivalents) in six different matrices to create 184 pools of BAC DNA. DNA fragments from each pool were amplified using amplified fragment length polymorphism (AFLP) technology, resolved on a LI-COR dual-dye DNA sequencing system, and analyzed using Bionumerics software. On average, each set of AFLP primers amplified 28 single-copy DNA markers that were useful for identifying overlapping BAC clones. Data from 32 different AFLP primer combinations identified ∼2400 BACs and ordered ∼700 BAC contigs. Analysis of a sorghum RIL mapping population using the same primer pairs located ∼200 of the BAC contigs on the sorghum genetic map. Restriction endonuclease fingerprinting of the entire collection of sorghum BAC clones was applied to test and extend the contigs constructed using this PCR-based methodology. Analysis of the fingerprint data allowed for the identification of 3366 contigs each containing an average of 5 BACs. BACs in ∼65% of the contigs aligned by AFLP analysis had sufficient overlap to be confirmed by DNA fingerprint analysis. In addition, 30% of the overlapping BACs aligned by AFLP analysis provided information for merging contigs and singletons that could not be joined using fingerprint data alone. Thus, the combination of fingerprinting and AFLP-based contig assembly and mapping provides a reliable, high-throughput method for building an integrated genetic and physical map of the sorghum genome.[The sequence data described in this paper have been submitted to the GenBank data library under accession no. AF218263.]Integrated genetic and physical genome maps are extremely valuable for map-based gene isolation, comparative genome analysis, and as sources of sequenceready clones for genome sequencing projects. Various methods have been developed for assembling physical maps of complex genomes. One of the best characterized approaches uses restriction enzymes to generate large numbers of DNA fragments from genomic subclones (Brenner and Livak 1989;Gregory et al. 1997;Marra et al. 1997). These DNA fingerprints are compared to identify related clones, and to assemble overlapping clones in contigs. The utility of fingerprinting for ordering a complex genome is limited, however, due to variation in DNA migration from gel to gel, the presence of repetitive DNAs, unusual distribution of restriction sites and skewed clone representation. Moreover, fingerprinting, unless combined with other methods, does not link genomic clones directly to gen...

Genome wide changes in gene expression were monitored in the drought tolerant C4 cereal Sorghum bicolor, following exposure of seedlings to high salinity (150 mM NaCl), osmotic stress (20% polyethylene glycol) or abscisic acid (125 microM ABA). A sorghum cDNA microarray providing data on 12,982 unique gene clusters was used to examine gene expression in roots and shoots at 3- and 27-h post-treatment. Expression of approximately 2200 genes, including 174 genes with currently unknown functions, of which a subset appear unique to monocots and/or sorghum, was altered in response to dehydration, high salinity or ABA. The modulated sorghum genes had homology to proteins involved in regulation, growth, transport, membrane/protein turnover/repair, metabolism, dehydration protection, reactive oxygen scavenging, and plant defense. Real-time PCR was used to quantify changes in relative mRNA abundance for 333 genes that responded to ABA, NaCl or osmotic stress. Osmotic stress inducible sorghum genes identified for the first time included a beta-expansin expressed in shoots, actin depolymerization factor, inositol-3-phosphate synthase, a non-C4 NADP-malic enzyme, oleosin, and three genes homologous to 9-cis-epoxycarotenoid dioxygenase that may be involved in ABA biosynthesis. Analysis of response profiles demonstrated the existence of a complex gene regulatory network that differentially modulates gene expression in a tissue- and kinetic-specific manner in response to ABA, high salinity and water deficit. Modulation of genes involved in signal transduction, chromatin structure, transcription, translation and RNA metabolism contributes to sorghum's overlapping but nonetheless distinct responses to ABA, high salinity, and osmotic stress. Overall, this study provides a foundation of information on sorghum's osmotic stress responsive gene complement that will accelerate follow up biochemical, QTL and comparative studies.

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

Background: Sorghum genome mapping based on DNA markers began in the early 1990s and numerous genetic linkage maps of sorghum have been published in the last decade, based initially on RFLP markers with more recent maps including AFLPs and SSRs and very recently, Diversity Array Technology (DArT) markers. It is essential to integrate the rapidly growing body of genetic linkage data produced through DArT with the multiple genetic linkage maps for sorghum generated through other marker technologies. Here, we report on the colinearity of six independent sorghum component maps and on the integration of these component maps into a single reference resource that contains commonly utilized SSRs, AFLPs, and high-throughput DArT markers.

The objectives of this study were (i) to assess the level of genetic diversity in elite sterility‐maintaining (B) and fertility‐restoring (R) sorghum [Sorghum bicolor (L.) Moench] lines as compared with a group of exotic and converted germplasm (IS) from the World Collection, (ii) to compare the classification of germplasm on the basis of estimates of genetic similarities obtained by means of AFLP and microsatellite (SSR) markers, and (iii) to compare the classification of germplasm obtained by different classes of molecular markers. A set of 100 SSRs, 1318 EcoRI/MseI AFLP, and 496 PstI/MseI AFLP markers with known map positions were utilized to determine the genetic similarity in a group of B, R, and IS public inbreds. Cluster analysis of genetic similarity estimates (GSij) revealed that the classification of sorghum inbreds is based on the sorghum working groups, Zera‐zera, Kafir, Kafir‐Milo, Durra, and Feterita. Cluster analyses failed to give a clear differentiation between B‐ and R‐lines, suggesting that R‐ and B‐lines do not represent well‐defined heterotic groups in this set of public lines. By comparing the different classes of molecular markers (SSRs, AFLPs, combinations of SSRs and AFLPs), we determined that the distribution of the markers and the coverage of the genome by the markers did affect the classification of genotypes. Dendrograms of genetic similarity (GS) based on PstI/MseI AFLP markers, or a set of markers spaced at 1‐ to 2‐cM intervals across the genome, produced clusters that were in better agreement with pedigree information than the analysis based solely on the EcoRI/MseI AFLP or SSR markers used in this study.