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Abstract Introduction This study was designed to analyze whether routine radiological controls of anastomoses in the upper gastrointestinal tract an early detection of anastomotic leaks. Patients and Methods 135 patients who underwent upper gastrointestinal tract surgery were retrospectively analyzed. Patients in the first group (n = 55) underwent routine radiological control of the anastomoses. In the second group (n = 80) the radiological control was only performed in case of clinical symptoms or signs of anastomotic leaks. Results The incidence of anastomotic leaks in the patients seen by us was 5.2%, equivalent to 7 of 135 patients In Group 1 leaks were seen in 4 of 55 patients (7,2%) in group 2 leaks were seen in 3 of 80 (3,8%). The radiological control of the anastomoses with contrast swallow showed the leakage in two cases. Twice the results were false negative. The sensitivity of computed tomography was 100%. Discussion Routine radiological control of anastomoses with contrast swallow only has low sensitivity. This procedure should not be performed routinely any more. The radiological control should be used in cases with signs of anastomotic leakage or with postoperatively impaired gastrointestinal passage.

Black pericarp sorghum has notable value due to the biosynthesis of 3-deoxyanthocyanidins (3-DOAs), a rare class of bioactive polyphenols valued as antioxidant food additives and as bioactive compounds with cytotoxicity to human cancer cells. A metabolic and transcriptomic study was conducted to ascertain the cellular events leading to the activation of 3-DOA biosynthesis in black sorghum pericarp. Prolonged exposure of pericarp during grain maturation to high-fluence ultraviolet (UV) light resulted in elevated levels of reactive oxygen species (ROS) and the activation of 3-DOA biosynthesis in pericarp tissues. In conjunction with 3-DOA biosynthesis was the transcriptional activation of specific family members of early and late flavonoid biosynthesis pathway genes as well as the downstream activation of defense-related pathways. Promoter analysis of genes highly correlated with 3-DOA biosynthesis in black pericarp were enriched in MYB and HHO5/ARR-B motifs. Light microscopy studies of black pericarp tissues suggest that 3-DOAs are predominantly localized in the epicarp and are associated with the cell wall. A working model of UV-induced 3-DOA biosynthesis in black pericarp is proposed that shares features of plant immunity associated with pathogen attack or mechanical wounding. The present model depicts ROS accumulation, the transcriptional activation of receptor kinases and transcription factors (TFs) including NAC, WRKY, bHLH, AP2, and C2H2 Zinc finger domain. This study identified key biosynthetic and regulatory genes of 3-DOA accumulation in black pericarp and provided a deeper understanding of the gene networks and cellular events controlling this tissue-and genotype-specific trait., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Schumaker, Mortensen, Klein, Mandal, Dykes, Gladman, Rooney, Burson and Klein.)

Introduction: Prickles are often deemed undesirable traits in many crops, including roses ( Rosa sp.), and there is demand for rose cultivars with no or very few prickles. This study aims to identify new and/or validate reported quantitative trait loci (QTLs) associated with stem and leaf rachis prickle density, characterize the effects of functional haplotypes for major QTLs, and identify the sources of QTL-alleles associated with increased/decreased prickle density in roses., Methods: QTL mapping using pedigree-based analysis (PBA), and haplotype analysis were conducted on two multi-parental diploid rose populations (TX2WOB and TX2WSE)., Results and Discussion: Twelve QTLs were identified on linkage groups (LGs) 2, 3, 4, and 6. The major QTLs for the stem prickle density were located between 42.25 and 45.66 Mbp on chromosome 3 of the Rosa chinensis genome assembly, with individual QTLs explaining 18 to 49% of phenotypic variance (PVE). The remaining mapped QTLs were minor. As for the rachis prickle density, several QTLs were detected on LG3, 4, and 6 with PVE 8 to 17%. Also, this study identified that ancestors R. wichurana 'Basye's Thornless', 'Old Blush', and the pollen parent of M4-4 were common sources of favorable alleles ( q ) associated with decreased prickle density, whereas 'Little Chief' and 'Srche Europy' were the source of unfavorable alleles ( Q ) in the TX2WOB and TX2WSE populations, respectively. The outcomes of this work complement other studies to locate factors that affect prickle density. These results can also be utilized to develop high-throughput DNA tests and apply parental selection to develop prickle-free rose cultivars., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Rawandoozi, Barocco, Rawandoozi, Klein, Byrne and Riera-Lizarazu.)

Maintaining or introducing genetic diversity into plant breeding programs is necessary for continual genetic gain; however, diversity at the cost of reduced performance is not something sought by breeders. To this end, backcross-nested association mapping (BC-NAM) populations, in which the recurrent parent is an elite line, can be employed as a strategy to introgress diversity from unadapted accessions while maintaining agronomic performance. This study evaluates (i) the hybrid performance of sorghum lines from 18 BC 1 -NAM families and (ii) the potential of genomic prediction to screen lines from BC 1 -NAM families for hybrid performance prior to phenotypic evaluation. Despite the diverse geographical origins and agronomic performance of the unadapted parents for BC 1 -NAM families, many BC 1 -derived lines performed significantly better in the hybrid trials than the elite recurrent parent, R.Tx436. The genomic prediction accuracies for grain yield, plant height, and days to mid-anthesis were acceptable, but the prediction accuracies for plant height were lower than expected. While the prediction accuracies increased when including more individuals in the training set, improvements tended to plateau between two and five lines per family, with larger training sets being required for more complex traits such as grain yield. Therefore, genomic prediction models can be optimized in a large BC 1 -NAM population with a relatively low fraction of individuals needing to be evaluated. These results suggest that genomic prediction is an effective method of pre-screening lines within BC 1 -NAM families prior to evaluation in extensive hybrid field trials.

Use of trifluoromethanesulfonamide (TFMSA), a male gametocide, increases the opportunities to identify promising B-lines because large quantities of F 1 seed can be generated prior to the laborious task of B-line sterilization. Combining TFMSA technology with genomic selection could efficiently evaluate sorghum B-lines in hybrid combination to maximize the rates of genetic gain of the crop. This study used two recombinant inbred B-line populations, consisting of 217 lines, which were testcrossed to two R-lines to produce 434 hybrids. Each population of testcross hybrids were evaluated across five environments. Population-based genomic prediction models were assessed across environments using three different cross-validation (CV) schemes, each with 70% training and 30% validation sets. The validation schemes were as follows: CV1-hybrids chosen randomly for validation; CV2-B-lines were randomly chosen, and each chosen B-line had one of the two corresponding testcross hybrids randomly chosen for the validation; and CV3-B-lines were randomly chosen, and each chosen B-line had both corresponding testcross hybrids chosen for the validation. CV1 and CV2 presented the highest prediction accuracies; nonetheless, the prediction accuracies of the CV schemes were not statistically different in many environments. We determined that combining the B-line populations could improve prediction accuracies, and the genomic prediction models were able to effectively rank the poorest 70% of hybrids even when genomic prediction accuracies themselves were low. Results indicate that combining genomic prediction models and TFMSA technology can effectively aid breeders in predicting B-line hybrid performance in early generations prior to the laborious task of generating A/B-line pairs., (© 2023 The Authors. The Plant Genome published by Wiley Periodicals LLC on behalf of Crop Science Society of America.)

Rose ( Rosa spp.) is one of the most economically important ornamental species worldwide. Flower diameter, flower weight, and the number of petals and petaloids are key flower-size parameters and attractive targets for DNA-informed breeding. Pedigree-based analysis (PBA) using FlexQTL software was conducted using two sets of multi-parental diploid rose populations. Phenotypic data for flower diameter (Diam), flower weight (fresh (FWT)/dry (DWT)), number of petals (NP), and number of petaloids (PD) were collected over six environments (seasons) at two locations in Texas. The objectives of this study were to 1) identify new and/or validate previously reported QTL(s); 2) identify SNP haplotypes associated with QTL alleles ( Q -/ q- ) of a trait and their sources; and 3) determine QTL genotypes for important rose breeding parents. Several new and previously reported QTLs for NP and Diam traits were identified. In addition, QTLs associated with flower weight and PD were identified for the first time. Two major QTLs with large effects were mapped for all traits. The first QTL was at the distal end of LG1 (60.44-60.95 Mbp) and was associated with Diam and DWT in the TX2WOB populations. The second QTL was consistently mapped in the middle region on LG3 (30.15-39.34 Mbp) and associated with NP, PD, and flower weight across two multi-parent populations (TX2WOB and TX2WSE). Haplotype results revealed a series of QTL alleles with differing effects at important loci for most traits. This work is distinct from previous studies by conducting co-factor analysis to account for the DOUBLE FLOWER locus while mapping QTL for NP. Sources of high-value ( Q ) alleles were identified, namely, 'Old Blush' and Rosa wichuraiana from J14-3 for Diam, while 'Violette' and PP-J14-3 were sources for other traits. In addition, the source of the low-value ( q ) alleles for Diam was 'Little Chief', and Rosa wichuraiana through J14-3 was the source for the remaining traits. Hence, our results can potentially inform parental/seedling selections as means to improve ornamental quality in roses and a step towards implementing DNA-informed techniques for use in rose breeding programs., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Rawandoozi, Young, Liang, Wu, Fu, Hochhaus, Yan, Rawandoozi, Klein, Byrne and Riera-Lizarazu.)

Garden roses are an economically important horticultural crop worldwide, and two major fungal pathogens, black spot ( Diplocarpon rosae F.A. Wolf) and cercospora leaf spot of rose ( Rosisphaerella rosicola Pass.), affect both the health and ornamental value of the plant. Most studies on black spot disease resistance have focused on diploid germplasm, and little work has been performed on cercospora leaf spot resistance. With the use of newly developed software tools for autopolyploid genetics, two interconnected tetraploid garden rose F 1 populations (phenotyped over the course of 3 years) were used for quantitative trait locus (QTL) analysis of black spot and cercospora leaf spot resistance as well as plant defoliation. QTLs for black spot resistance were mapped to linkage groups (LGs) 1-6. QTLs for cercospora resistance and susceptibility were found in LGs 1, 4, and 5 and for defoliation in LGs 1, 3, and 5. The major locus on LG 5 for black spot resistance coincides with the previously discovered Rdr4 locus inherited from Rosa L. 'Radbrite' (Brite Eyes™), the common parent used in these mapping populations. This work is the first report of any QTL for cercospora resistance/susceptibility in tetraploid rose germplasm and the first report of defoliation QTL in roses. A major QTL for cercospora susceptibility coincides with the black spot resistance QTL on LG 5 ( Rdr4 ). A major cercospora resistance QTL was found on LG 1. These populations provide a genetic resource that will further the knowledge base of rose genetics as more traits are studied. Studying more traits from these populations will allow for the stacking of various QTLs for desirable traits., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Lau, Gill, Taniguti, Young, Klein, Byrne and Riera-Lizarazu.)

To exploit the novel genetic diversity residing in tropical sorghum germplasm, an expansive backcross nested-association mapping (BC-NAM) resource was developed in which novel genetic diversity was introgressed into elite inbreds. A major limitation of exploiting this type of genetic resource in hybrid improvement programs is the required evaluation in hybrid combination of the vast number of BC-NAM populations and lines. To address this, the utility of genomic information was evaluated to predict the hybrid performance of BC-NAM populations. Two agronomically elite BC-NAM populations were chosen for evaluation in which elite inbred RTx436 was the recurrent parent. Each BC 1 F 3 line was evaluated in hybrid combination with an elite tester in two locations with phenotypes of grain yield, plant height, and days to anthesis collected on all test cross hybrids. Lines from both populations were found to outperform their recurrent parent. Efforts to utilize genetic distance based on genotyping-by-sequence (GBS) as a predictive tool for hybrid performance was ineffective. However, utilizing genomic prediction models using additive and dominance GBLUP kernels to screen germplasm appeared to be an effective method to eliminate inferior-performing lines that will not be useful in a hybrid breeding program.

Cercospora leaf spot (CLS) ( Cercospora rosicola ) is a major fungal disease of roses ( Rosa sp.) in the southeastern U.S. Developing CLS-resistant cultivars offers a potential solution to reduce pesticide use. Yet, no work has been performed on CLS resistance. This study aimed to identify QTLs and to characterize alleles for resistance to CLS. The study used pedigree-based QTL analysis to dissect the genetic basis of CLS resistance using two multi-parental diploid rose populations (TX2WOB and TX2WSE) evaluated across five years in two Texas locations. A total 38 QTLs were identified across both populations and distributed over all linkage groups. Three QTLs on LG3, LG4, and LG6 were consistently mapped over multiple environments. The LG3 QTL was mapped in a region between 18.9 and 27.8 Mbp on the Rosa chinensis genome assembly. This QTL explained 13 to 25% of phenotypic variance. The LG4 QTL detected in the TX2WOB population spanned a 35.2 to 39.7 Mbp region with phenotypic variance explained (PVE) up to 48%. The LG6 QTL detected in the TX2WSE population was localized to 17.9 to 33.6 Mbp interval with PVE up to 36%. Also, this study found multiple degrees of favorable allele effects ( q -allele) associated with decreasing CLS at major loci. Ancestors 'OB', 'Violette', and PP-M4-4 were sources of resistance q -alleles. These results will aid breeders in parental selection to develop CLS-resistant rose cultivars. Ultimately, high throughput DNA tests that target major loci for CLS could be developed for routine use in a DNA-informed breeding program., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2023 Rawandoozi, Young, Kang, Yan, Noyan, Fu, Hochhaus, Rawandoozi, Klein, Byrne and Riera-Lizarazu.)

The stay-green trait is recognized as a key drought adaptation mechanism in cereals worldwide. Stay-green sorghum plants exhibit delayed senescence of leaves and stems, leading to prolonged growth, a reduced risk of lodging, and higher grain yield under end-of-season drought stress. More than 45 quantitative trait loci (QTL) associated with stay-green have been identified, including two major QTL (Stg1 and Stg2). However, the contributing genes that regulate functional stay-green are not known. Here we show that the PIN FORMED family of auxin efflux carrier genes induce some of the causal mechanisms driving the stay-green phenotype in sorghum, with SbPIN4 and SbPIN2 located in Stg1 and Stg2, respectively. We found that nine of 11 sorghum PIN genes aligned with known stay-green QTL. In transgenic studies, we demonstrated that PIN genes located within the Stg1 (SbPIN4), Stg2 (SbPIN2), and Stg3b (SbPIN1) QTL regions acted pleiotropically to modulate canopy development, root architecture, and panicle growth in sorghum, with SbPIN1, SbPIN2, and SbPIN4 differentially expressed in various organs relative to the non-stay-green control. The emergent consequence of such modifications in canopy and root architecture is a stay-green phenotype. Crop simulation modelling shows that the SbPIN2 phenotype can increase grain yield under drought., (© The Author(s) 2022. Published by Oxford University Press on behalf of the Society for Experimental Biology.)

Black spot disease (BSD) ( Diplocarpon rosae ) is the most common and damaging fungal disease in garden roses ( Rosa sp .). Although qualitative resistance to BSD has been extensively investigated, the research on quantitative resistance lags behind. The goal of this research was to study the genetic basis of BSD resistance in two multi-parental populations (TX2WOB and TX2WSE) through a pedigree-based analysis approach (PBA). Both populations were genotyped and evaluated for BSD incidence over five years in three locations in Texas. A total of 28 QTLs, distributed over all linkage groups (LGs), were detected across both populations. Consistent minor effect QTLs included two on LG1 and LG3 (TX2WOB and TX2WSE), two on LG4 and LG5 (TX2WSE), and one QTL on LG7 (TX2WOB). In addition, one major QTL detected in both populations was consistently mapped on LG3. This QTL was localized to an interval ranging from 18.9 to 27.8 Mbp on the Rosa chinensis genome and explained 20 and 33% of the phenotypic variation. Furthermore, haplotype analysis showed that this QTL had three distinct functional alleles. The parent PP-J14-3 was the common source of the LG3 BSD resistance in both populations. Taken together, this research presents the characterization of new SNP-tagged genetic determinants of BSD resistance, the discovery of marker-trait associations to enable parental choice based on their BSD resistance QTL haplotypes, and substrates for the development of trait-predictive DNA tests for routine use in marker-assisted breeding for BSD resistance., Competing Interests: The authors declare there are no conflicts of interest., (© The Author(s) 2022. Published by Oxford University Press on behalf of Nanjing Agricultural University.)

Rose rosette disease (RRD), caused by the Rose rosette emaravirus (RRV), is a major threat to the garden rose industry in the United States. There has been limited work on the genetics of host plant resistance to RRV. Two interconnected tetraploid garden rose F 1 biparental mapping populations were created to develop high-quality tetraploid rose linkage maps that allowed the discovery of RRD resistance quantitative trait loci (QTLs) on linkage groups (LGs) 5, 6, and 7. These QTLs individually accounted for around 18-40% of the phenotypic variance. The locus with the greatest effect on partial resistance was found in LG 5. Most individuals with the LG 5 QTL were in the simplex configuration; however, two individuals were duplex (likely due to double reduction). Identification of resistant individuals and regions of interest can help the development of diagnostic markers for marker-assisted selection in a breeding program., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2022 Lau, Young, Collins, Windham, Klein, Byrne and Riera-Lizarazu.)

Resistance to rose rosette disease (RRD), a fatal disease of roses ( Rosa spp.), is a high priority for rose breeding. As RRD resistance is time-consuming to phenotype, the identification of genetic markers for resistance could expedite breeding efforts. However, little is known about the genetics of RRD resistance. Therefore, we performed a quantitative trait locus (QTL) analysis on a set of inter-related diploid rose populations phenotyped for RRD resistance and identified four QTLs. Two QTLs were found in multiple years. The most consistent QTL is qRRV_TX2WSE_ch5 , which explains approximately 20% and 40% of the phenotypic variation in virus quantity and severity of RRD symptoms, respectively. The second, a QTL on chromosome 1, qRRD_TX2WSE_ch1 , accounts for approximately 16% of the phenotypic variation for severity. Finally, a third QTL on chromosome 3 was identified only in the multiyear analysis, and a fourth on chromosome 6 was identified in data from one year only. In addition, haplotypes associated with significant changes in virus quantity and severity were identified for qRRV_TX2WSE_ch5 and qRRD_TX2WSE_ch1 . This research represents the first report of genetic determinants of resistance to RRD. In addition, marker trait associations discovered here will enable better parental selection when breeding for RRD resistance and pave the way for marker-assisted selection for RRD resistance.

Genomic selection in maize (Zea mays L.) has been one factor that has increased the rate of genetic gain when compared with other cereals. However, the technological foundations in maize also exist in other cereal crops that would allow prediction of hybrid performance based on general (GCA) and specific (SCA) combining abilities applied through genomic-enabled prediction models. Further, the incorporation of genotype × environment (G × E) interaction effects present an opportunity to deploy hybrids to targeted environments. To test these concepts, a factorial mating design of elite yet divergent grain sorghum lines generated hybrids for evaluation. Inbred parents were genotyped, and markers were used to assess population structure and develop the genomic relationship matrix (GRM). Grain yield, height, and days to anthesis were collected for hybrids in replicated trials, and best linear unbiased estimates were used to train classical GCA-SCA-based and genomic (GB) models under a hierarchical Bayesian framework. To incorporate population structure, GB was fitted using the GRM of both parents and hybrids. For GB models, G × E interaction effects were included by the Hadamard product between GRM and environments. A leave-one-out cross-validation scheme was used to study the prediction capacity of models. Classical and genomic models effectively predicted hybrid performance and prediction accuracy increased by including genomic data. Genomic models effectively partitioned the variation due to GCA, SCA, and their interaction with the environment. A strategy to implement genomic selection for hybrid sorghum [Sorghum bicolor (L.) Moench] breeding is presented herein., (© 2021 The Authors. The Plant Genome published by Wiley Periodicals LLC on behalf of Crop Science Society of America.)

Genome-enabled biotechnologies have the potential to accelerate breeding efforts in long-lived perennial crop species. Despite the transformative potential of molecular tools in pecan and other outcrossing tree species, highly heterozygous genomes, significant presence-absence gene content variation, and histories of interspecific hybridization have constrained breeding efforts. To overcome these challenges, here, we present diploid genome assemblies and annotations of four outbred pecan genotypes, including a PacBio HiFi chromosome-scale assembly of both haplotypes of the 'Pawnee' cultivar. Comparative analysis and pan-genome integration reveal substantial and likely adaptive interspecific genomic introgressions, including an over-retained haplotype introgressed from bitternut hickory into pecan breeding pedigrees. Further, by leveraging our pan-genome presence-absence and functional annotation database among genomes and within the two outbred haplotypes of the 'Lakota' genome, we identify candidate genes for pest and pathogen resistance. Combined, these analyses and resources highlight significant progress towards functional and quantitative genomics in highly diverse and outbred crops.

Black sorghum [ Sorghum bicolor (L.) Moench] is characterized by the black appearance of the pericarp and production of 3-deoxyanthocyanidins (3-DOA), which are valued for their cytotoxicity to cancer cells and as natural food colorants and antioxidant additives. The black pericarp phenotype is not fully penetrant in all environments, which implicates the light spectrum and/or photoperiod as the critical factor for trait expression. In this study, black- or red-pericarp genotypes were grown under regimes of visible light, visible light supplemented with UVA or supplemented with UVA plus UVB (or dark control). Pericarp 3-DOAs and pericarp pigmentation were maximized in the black genotype exposed to a light regime supplemented with UVB. Changes in gene expression during black pericarp development revealed that ultraviolet light activates genes related to plant defense, reactive oxygen species, and secondary metabolism, suggesting that 3-DOA accumulation is associated with activation of flavonoid biosynthesis and several overlapping defense and stress signaling pathways.

The projected impact of global warming on coffee production may require the heat-adapted genotypes in the next decades. To identify cellular strategies in response to warmer temperatures, we compared the effect of elevated temperature on two commercial Coffea arabica L. genotypes exploring leaf physiology, transcriptome, and carbohydrate/protein composition. Growth temperatures were 23/19°C (day/night), as optimal condition (OpT), and 30/26°C (day/night) as a possible warmer scenario (WaT). The cv. Acauã showed lower levels of leaf temperature (Tleaf) under both conditions compared to cv. Catuaí, whereas slightly or no differences for other leaf physiological parameters. Therefore, to explore temperature responsive pathways the leaf transcriptome was examined using RNAseq. Genotypes showed a marked number of differentially-expressed genes (DEGs) under OpT, however DEGs strongly decrease in both at WaT condition indicating a transcriptional constraint. DEGs responsive to WaT revealed shared and genotype-specific genes mostly related to carbohydrate metabolism. Under OpT, leaf starch content was greater in cv. Acauã and, as WaT temperature was imposed, the leaf soluble sugar did not change in contrast to cv. Catuaí, although the levels of leaf starch, sucrose, and leaf protein decreased in both genotypes. These findings revealed intraspecific differences in the underlying transcriptional and metabolic interconnected pathways responsive to warmer temperatures, which is potentially linked to thermotolerance, and thus may be useful as biomarkers in breeding for a changing climate., (Copyright © 2020 de Oliveira, Ribeiro, Cardon, Fedenia, Maia, Barbosa, Caldeira, Klein and Chalfun-Junior.)

The genome of the allotetraploid species Coffea arabica L. was sequenced to assemble independently the two component subgenomes (putatively deriving from C. canephora and C. eugenioides) and to perform a genome-wide analysis of the genetic diversity in cultivated coffee germplasm and in wild populations growing in the center of origin of the species. We assembled a total length of 1.536 Gbp, 444 Mb and 527 Mb of which were assigned to the canephora and eugenioides subgenomes, respectively, and predicted 46,562 gene models, 21,254 and 22,888 of which were assigned to the canephora and to the eugeniodes subgenome, respectively. Through a genome-wide SNP genotyping of 736 C. arabica accessions, we analyzed the genetic diversity in the species and its relationship with geographic distribution and historical records. We observed a weak population structure due to low-frequency derived alleles and highly negative values of Taijma's D, suggesting a recent and severe bottleneck, most likely resulting from a single event of polyploidization, not only for the cultivated germplasm but also for the entire species. This conclusion is strongly supported by forward simulations of mutation accumulation. However, PCA revealed a cline of genetic diversity reflecting a west-to-east geographical distribution from the center of origin in East Africa to the Arabian Peninsula. The extremely low levels of variation observed in the species, as a consequence of the polyploidization event, make the exploitation of diversity within the species for breeding purposes less interesting than in most crop species and stress the need for introgression of new variability from the diploid progenitors.

Multiparent advanced generation inter-cross (MAGIC) populations can provide improved genetic mapping resolution by increasing allelic diversity and effective recombination. The Four Parent Maize (FPM; L.) population implemented five different mating designs used in MAGIC and bi-parental populations to compare empirical effects on genetic resolution and power of quantitative trait locus (QTL) detection; the combined population here comprised of 1149 individuals with 118,509 genetic markers. Measurements were recorded for plant height (PH), ear height (EH), days to anthesis (DTA) and silking (DTS) in seven environments, spanning three years. Linkage disequilibrium (LD) analysis of subpopulations indicated MAGIC population designs should incorporate generations of intermating to overcome initial LD increase caused by population admixture in a non-intermated four parent population (4way0sib). A 3- to 4-fold increase in genetic resolution (<0.8) and a 2.5-fold decrease in the extent of LD decay (<0.2) compared to the biparental populations was found for the four parent cross at the third generation of intermating (4way3sib). Power of QTL detection was affected to a greater extent by sample size rather than by mating designs. The FPM power simulations indicated that MAGIC populations have the ability to meet or exceed the mapping power of nested association panels with fewer individuals and diversity inputs. Using association mapping software we identified 2, 5, 7, and 6 QTL for PH, EH, DTA, and DTS, respectively. The FPM population is a valuable resource for quantifying empirical improvements of parent number, intermating, and the number of progeny for QTL linkage mapping., (Copyright © 2018 Crop Science Society of America.)

Roses, which have been cultivated for at least 5000 years, are one of the most important ornamental crops in the world. Because of the interspecific nature and high heterozygosity in commercial roses, the genetic resources available for rose are limited. To effectively identify markers associated with QTL controlling important traits, such as disease resistance, abundant markers along the genome and careful phenotyping are required. Utilizing genotyping by sequencing technology and the strawberry genome ( Fragaria vesca v2.0.a1) as a reference, we generated thousands of informative single nucleotide polymorphism (SNP) markers. These SNPs along with known bridge simple sequence repeat (SSR) markers allowed us to create the first high-density integrated consensus map for diploid roses. Individual maps were first created for populations J06-20-14-3×"Little Chief" (J14-3×LC), J06-20-14-3×"Vineyard Song" (J14-3×VS) and "Old Blush"×"Red Fairy" (OB×RF) and these maps were linked with 824 SNPs and 13 SSR bridge markers. The anchor SSR markers were used to determine the numbering of the rose linkage groups. The diploid consensus map has seven linkage groups (LGs), a total length of 892.2 cM, and an average distance of 0.25 cM between 3527 markers. By combining three individual populations, the marker density and the reliability of the marker order in the consensus map was improved over a single population map. Extensive synteny between the strawberry and diploid rose genomes was observed. This consensus map will serve as the tool for the discovery of marker-trait associations in rose breeding using pedigree-based analysis. The high level of conservation observed between the strawberry and rose genomes will help further comparative studies within the Rosaceae family and may aid in the identification of candidate genes within QTL regions., Competing Interests: The authors declare that they have no conflict of interest.

'Jones Hybrid' hickory is an accession in the National Collection of Genetic Resources for Pecans and Hickories for which information about origin, identity and characteristics is very incomplete. Phenotypic and genetic profiles, when examined in the context of historic literature, provide evidence that the accession in question is 'Siers' a cultivar of Carya  ×  laneyi (an interspecific hybrid between C. ovata and C. cordiformis ). The accession has traits that make it interesting in the pecan breeding program, with potential for both rootstock and scion development. The tall, slender tree form of 'Jones Hybrid' is a trait that could be valuable in commercial pecan cultivars, allowing increased tree densities and reducing the need for expensive hedging operations. Tree size reduction is a goal to be pursued in scion selection and rootstock development, with each goal requiring assessment of reproductive potential of the accession.

Sorghum is one of the world's most important food, feed, and fiber crops as well as a potential feedstock for lignocellulosic bioenergy. Early-season planting extends sorghum's growing season and increases yield in temperate regions. However, sorghum's sensitivity to low soil temperatures adversely impacts seed germination. In this study, we evaluated the 242 accessions of the ICRISAT sorghum mini core collection for seed germination and seedling vigor at 12 °C as a measure of cold tolerance. Genome-wide association analysis was performed with approximately 162,177 single nucleotide polymorphism markers. Only one marker locus (Locus 7-2) was significantly associated with low-temperature germination and none with vigor. The linkage of Locus 7-2 to low-temperature germination was supported by four lines of evidence: strong association in three independent experiments, co-localization with previously mapped cold tolerance quantitative trait loci (QTL) in sorghum, a candidate gene that increases cold tolerance and germination rate when its wheat homolog is overexpressed in tobacco, and its syntenic region in rice co-localized with two cold tolerance QTL in rice. This locus may be useful in developing tools for molecular breeding of sorghums with improved low-temperature germinability.

Key Message: Allele phylogenetic analysis of the sorghum flowering-time gene PRR37 provided new insight into the human-mediated selection of a key adaptive gene that occurred during sorghum's diversification and worldwide dispersal. The domestication and spread of the tropical cereal sorghum is associated with the historic movement of humans. We show that an allelic series at PRR37 (pseudo-response regulator 37), a circadian clock-associated transcription factor, was selected in long-day ecosystems worldwide to permit floral initiation and grain production. We identified a series of loss-of-function (photoperiod-insensitive) alleles encoding truncated PRR37 proteins, alleles with key amino acid substitutions in the pseudo-receiver domain, and a novel splice variant in which the pseudo-receiver domain is truncated. Each PRR37 allelic variant was traced to a specific geographic location or specialized agronomic type. We present a graphical model that shows evidence of human selection and gene flow of the PRR37 allelic variants during the global dispersal and agronomic diversification of sorghum. With the recent identification of the Ghd7 gene as an important regulator of flowering date in sorghum, we briefly examine whether loss-of-function Ghd7 allelic variants were selected prior to the human-mediated movement of sorghum from its equatorial center of origin to temperate climates worldwide.

Imperata cylindrica (Cogongrass, Speargrass) is a diploid C4 grass that is a noxious weed in 73 countries and constitutes a significant threat to global biodiversity and sustainable agriculture. We used a cost-effective genotyping-by-sequencing (GBS) approach to identify the reproductive system, genetic diversity and geographic origins of invasions in the south-eastern United States. In this work, we demonstrated the advantage of employing the closely related, fully sequenced crop species Sorghum bicolor (L.) Moench as a proxy reference genome to identify a set of 2320 informative single nucleotide and insertion-deletion polymorphisms. Genetic analyses identified four clonal lineages of cogongrass and one clonal lineage of Imperata brasiliensis Trin. in the United States. Each lineage was highly homogeneous, and we found no evidence of hybridization among the different lineages, despite geographical overlap. We found evidence that at least three of these lineages showed clonal reproduction prior to introduction to the United States. These results indicate that cogongrass has limited evolutionary potential to adapt to novel environments and further suggest that upon arrival to its invaded range, this species did not require local adaptation through hybridization/introgression or selection of favourable alleles from a broad genetic base. Thus, cogongrass presents a clear case of broad invasive success, across a diversity of environments, in a clonal organism with limited genetic diversity., (© 2015 John Wiley & Sons Ltd.)

Stay-green sorghum plants exhibit greener leaves and stems during the grain-filling period under water-limited conditions compared with their senescent counterparts, resulting in increased grain yield, grain mass, and lodging resistance. Stay-green has been mapped to a number of key chromosomal regions, including Stg1, Stg2, Stg3, and Stg4, but the functions of these individual quantitative trait loci (QTLs) remain unclear. The objective of this study was to show how positive effects of Stg QTLs on grain yield under drought can be explained as emergent consequences of their effects on temporal and spatial water-use patterns that result from changes in leaf-area dynamics. A set of four Stg near-isogenic lines (NILs) and their recurrent parent were grown in a range of field and semicontrolled experiments in southeast Queensland, Australia. These studies showed that the four Stg QTLs regulate canopy size by: (1) reducing tillering via increased size of lower leaves, (2) constraining the size of the upper leaves; and (3) in some cases, decreasing the number of leaves per culm. In addition, they variously affect leaf anatomy and root growth. The multiple pathways by which Stg QTLs modulate canopy development can result in considerable developmental plasticity. The reduction in canopy size associated with Stg QTLs reduced pre-flowering water demand, thereby increasing water availability during grain filling and, ultimately, grain yield. The generic physiological mechanisms underlying the stay-green trait suggest that similar Stg QTLs could enhance post-anthesis drought adaptation in other major cereals such as maize, wheat, and rice., (© The Author 2014. Published by Oxford University Press on behalf of the Society for Experimental Biology.)

Light signaling by phytochrome B in long days inhibits flowering in sorghum by increasing expression of the long day floral repressors PSEUDORESPONSE REGULATOR PROTEIN (SbPRR37, Ma1) and GRAIN NUMBER, PLANT HEIGHT AND HEADING DATE 7 (SbGHD7, Ma6). SbPRR37 and SbGHD7 RNA abundance peaks in the morning and in the evening of long days through coordinate regulation by light and output from the circadian clock. 58 M, a phytochrome B deficient (phyB-1, ma3R) genotype, flowered ∼60 days earlier than 100 M (PHYB, Ma3) in long days and ∼11 days earlier in short days. Populations derived from 58 M (Ma1, ma3R, Ma5, ma6) and R.07007 (Ma1, Ma3, ma5, Ma6) varied in flowering time due to QTL aligned to PHYB/phyB-1 (Ma3), Ma5, and GHD7/ghd7-1 (Ma6). PHYC was proposed as a candidate gene for Ma5 based on alignment and allelic variation. PHYB and Ma5 (PHYC) were epistatic to Ma1 and Ma6 and progeny recessive for either gene flowered early in long days. Light signaling mediated by PhyB was required for high expression of the floral repressors SbPRR37 and SbGHD7 during the evening of long days. In 100 M (PHYB) the floral activators SbEHD1, SbCN8 and SbCN12 were repressed in long days and de-repressed in short days. In 58 M (phyB-1) these genes were highly expressed in long and short days. Furthermore, SbCN15, the ortholog of rice Hd3a (FT), is expressed at low levels in 100 M but at high levels in 58 M (phyB-1) regardless of day length, indicating that PhyB regulation of SbCN15 expression may modify flowering time in a photoperiod-insensitive manner.

Stay-green is an integrated drought adaptation trait characterized by a distinct green leaf phenotype during grain filling under terminal drought. We used sorghum (Sorghum bicolor), a repository of drought adaptation mechanisms, to elucidate the physiological and genetic mechanisms underpinning stay-green. Near-isogenic sorghum lines (cv RTx7000) were characterized in a series of field and managed-environment trials (seven experiments and 14 environments) to determine the influence of four individual stay-green (Stg1-4) quantitative trait loci (QTLs) on canopy development, water use and grain yield under post-anthesis drought. The Stg QTL decreased tillering and the size of upper leaves, which reduced canopy size at anthesis. This reduction in transpirational leaf area conserved soil water before anthesis for use during grain filling. Increased water uptake during grain filling of Stg near-isogenic lines (NILs) relative to RTx7000 resulted in higher post-anthesis biomass production, grain number and yield. Importantly, there was no consistent yield penalty associated with the Stg QTL in the irrigated control. These results establish a link between the role of the Stg QTL in modifying canopy development and the subsequent impact on crop water use patterns and grain yield under terminal drought., (© 2014 The Authors. New Phytologist © 2014 New Phytologist Trust.)

Sorghum (Sorghum bicolor (L.) Moench) is a high-yielding, stress tolerant energy crop for lignocellulosic-based biofuel production. Saccharification is a process by which hydrolytic enzymes break down lignocellulosic materials to fermentable sugars for biofuel production, and mapping and identifying genes underlying saccharification yield is an important first step to genetically improve the plant for higher biofuel productivity. In this study, we used the ICRISAT sorghum mini core germplasm collection and 14 739 single nucleotide polymorphism markers to map saccharification yield. Seven marker loci were associated with saccharification yield and five of these loci were syntenic with regions in the maize genome that contain quantitative trait loci underlying saccharification yield and cell wall component traits. Candidate genes from the seven loci were identified but must be validated, with the most promising candidates being β-tubulin, which determines the orientation of cellulose microfibrils in plant secondary cell walls, and NST1, a master transcription factor controlling secondary cell wall biosynthesis in fibers. Other candidate genes underlying the different saccharification loci included genes that play a role in vascular development and suberin deposition in plants. The identified loci and candidate genes provide information into the factors controlling saccharification yield and may facilitate increasing biofuel production in sorghum.

Background: Rapid acquisition of accurate genotyping information is essential for all genetic marker-based studies. For species with relatively small genomes, complete genome resequencing is a feasible approach for genotyping; however, for species with large and highly repetitive genomes, the acquisition of whole genome sequences for the purpose of genotyping is still relatively inefficient and too expensive to be carried out on a high-throughput basis. Sorghum bicolor is a C4 grass with a sequenced genome size of ~730 Mb, of which ~80% is highly repetitive. We have developed a restriction enzyme targeted genome resequencing method for genetic analysis, termed Digital Genotyping (DG), to be applied to sorghum and other grass species with large repeat-rich genomes., Results: DG templates are generated using one of three methylation sensitive restriction enzymes that recognize a nested set of 4, 6 or 8 bp GC-rich sequences, enabling varying depth of analysis and integration of results among assays. Variation in sequencing efficiency among DG markers was correlated with template GC-content and length. The expected DG allele sequence was obtained 97.3% of the time with a ratio of expected to alternative allele sequence acquisition of >20:1. A genetic map aligned to the sorghum genome sequence with an average resolution of 1.47 cM was constructed using 1,772 DG markers from 137 recombinant inbred lines. The DG map enhanced the detection of QTL for variation in plant height and precisely aligned QTL such as Dw3 to underlying genes/alleles. Higher-resolution NgoMIV-based DG haplotypes were used to trace the origin of DNA on SBI-06, spanning Ma1 and Dw2 from progenitors to BTx623 and IS3620C. DG marker analysis identified the correct location of two miss-assembled regions and located seven super contigs in the sorghum reference genome sequence., Conclusion: DG technology provides a cost-effective approach to rapidly generate accurate genotyping data in sorghum. Currently, data derived from DG are used for many marker-based analyses, including marker-assisted breeding, pedigree and QTL analysis, genetic map construction, map-based gene cloning and association studies. DG in combination with whole genome resequencing is dramatically accelerating all aspects of genetic analysis of sorghum, an important genetic reference for C4 grass species.

To facilitate the mapping of genes in sorghum [Sorghum bicolor (L.) Moench] underlying economically important traits, we analyzed the genetic structure and linkage disequilibrium in a sorghum mini core collection of 242 landraces with 13,390 single-nucleotide polymorphims. The single-nucleotide polymorphisms were produced using a highly multiplexed genotyping-by-sequencing methodology. Genetic structure was established using principal component, Neighbor-Joining phylogenetic, and Bayesian cluster analyses. These analyses indicated that the mini-core collection was structured along both geographic origin and sorghum race classification. Examples of the former were accessions from Southern Africa, East Asia, and Yemen. Examples of the latter were caudatums with widespread geographical distribution, durras from India, and guineas from West Africa. Race bicolor, the most primitive and the least clearly defined sorghum race, clustered among other races and formed only one clear bicolor-centric cluster. Genome-wide linkage disequilibrium analyses showed linkage disequilibrium decayed, on average, within 10-30 kb, whereas the short arm of SBI-06 contained a linkage disequilibrium block of 20.33 Mb, confirming a previous report of low recombination on this chromosome arm. Four smaller but equally significant linkage disequilibrium blocks of 3.5-35.5 kb were detected on chromosomes 1, 2, 9, and 10. We examined the genes encoded within each block to provide a first look at candidates such as homologs of GS3 and FT that may indicate a selective sweep during sorghum domestication.

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.

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.

With an aim to further characterize the cytoplasmic male sterility-fertility restoration system in sorghum, a major fertility restoration gene was mapped along with a second locus capable of partial restoration of pollen fertility. The major fertility restoration gene, Rf(5), was located on sorghum chromosome SBI-05, and was capable of restoring pollen fertility in both A(1) and A(2) male sterile cytoplasms. Depending on the restorer parent, mapping populations exhibited fertility restoration phenotypes that ranged from nearly bimodal distribution due to the action of Rf(5), to a more normalized distribution reflecting the action of Rf(5) and additional modifier/partial restoration genes. A second fertility restoration locus capable of partially restoring pollen fertility in A(1) cytoplasm was localized to chromosome SBI-04. Unlike Rf(5), this modifier/partial restorer gene acting alone resulted in less than 10% seed set in both A(1) and A(2) cytoplasms, and modified the extent of restoration conditioned by the major restorer Rf(5) in A(1) cytoplasm. In examining the genomic regions spanning the Rf(5) locus, a cluster of pentatricopeptide gene family members with high homology to rice Rf (1) and sorghum Rf (2) were identified as potential candidates encoding Rf(5).

Sorghum has been improved by public and private breeding programs utilizing germplasm mostly from within the species Sorghum bicolor. Recently, hybridization with an Australian species, S. macrospermum (AAB1B1YYZZ), has been demonstrated and the genomic relationship to S. bicolor (AAB1B1) shown to be partially compatible. For this species to be potentially useful to sorghum improvement programs, there must be documented introgression into an S. bicolor background. Fifteen BC1F1 progeny were recovered using the interspecific hybrid as a female and embryo rescue. In these progeny, chromosome numbers ranged from 35 to 70 and all were essentially male-sterile. Repeated backcrossing with S. bicolor pollen produced BC2F1 seed on 3 of the 15 BC1F1 plants. BC2F1 progeny had varying levels of male fertility; selfed seed set ranged from 0% to 95%, with only 2 individuals being completely male-sterile. Using AFLP and SSR markers, genomic introgression of S. macrospermum ranged from 0% to 18.6%. Cytogenetic analysis of 19 individuals revealed that chromosome numbers were 20, except for a single backcross that had 21 chromosomes. Molecular cytogenetic analysis confirmed the presence of recombinant introgression chromosomes as well as alien addition and alien substitution chromosomes within the BC2F1s.

The A1 cytoplasmic-nuclear male sterility system in sorghum is used almost exclusively for the production of commercial hybrid seed and thus, the dominant genes that restore male fertility in F(1) hybrids are of critical importance to commercial seed production. The genetics of fertility restoration in sorghum can appear complex, being controlled by at least two major genes with additional modifiers and additional gene-environment interaction. To elucidate the molecular processes controlling fertility restoration and to develop a marker screening system for this important trait, two sorghum recombinant inbred line populations were created by crossing a restorer and a non-restoring inbred line, with fertility phenotypes evaluated in hybrid combination with three unique cytoplasmic male sterile lines. In both populations, a single major gene segregated for restoration which was localized to chromosome SBI-02 at approximately 0.5 cM from microsatellite marker, Xtxp304. In the two populations we observed that approximately 85 and 87% of the phenotypic variation in seed set was associated with the major Rf gene on SBI-02. Some evidence for modifier genes was also observed since a continuum of partial restored fertility was exhibited by lines in both RIL populations. With the prior report (Klein et al. in Theor Appl Genet 111:994-1012, 2005) of the cloning of the major fertility restoration gene Rf1 in sorghum, the major fertility restorer locus identified in this study was designated Rf2. A fine-mapping population was used to resolve the Rf2 locus to a 236,219-bp region of chromosome SBI-02, which spanned ~31 predicted open reading frames including a pentatricopeptide repeat (PPR) gene family member. The PPR gene displayed high homology with rice Rf1. Progress towards the development of a marker-assisted screen for fertility restoration is discussed.

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., Results: The six component maps were constructed using the MultiPoint software. The lengths of the resulting maps varied between 910 and 1528 cM. The order of the 498 markers that segregated in more than one population was highly consistent between the six individual mapping data sets. The framework consensus map was constructed using a "Neighbours" approach and contained 251 integrated bridge markers on the 10 sorghum chromosomes spanning 1355.4 cM with an average density of one marker every 5.4 cM, and were used for the projection of the remaining markers. In total, the sorghum consensus map consisted of a total of 1997 markers mapped to 2029 unique loci (1190 DArT loci and 839 other loci) spanning 1603.5 cM and with an average marker density of 1 marker/0.79 cM. In addition, 35 multicopy markers were identified. On average, each chromosome on the consensus map contained 203 markers of which 58.6% were DArT markers. Non-random patterns of DNA marker distribution were observed, with some clear marker-dense regions and some marker-rare regions., Conclusion: The final consensus map has allowed us to map a larger number of markers than possible in any individual map, to obtain a more complete coverage of the sorghum genome and to fill a number of gaps on individual maps. In addition to overall general consistency of marker order across individual component maps, good agreement in overall distances between common marker pairs across the component maps used in this study was determined, using a difference ratio calculation. The obtained consensus map can be used as a reference resource for genetic studies in different genetic backgrounds, in addition to providing a framework for transferring genetic information between different marker technologies and for integrating DArT markers with other genomic resources. DArT markers represent an affordable, high throughput marker system with great utility in molecular breeding programs, especially in crops such as sorghum where SNP arrays are not publicly available.

Sorghum has been improved by public and private breeding programs utilizing germplasm mostly from within the species Sorghum bicolor. Until recently, cross-incompatibilities have prevented hybridization of S. bicolor with most other species within the genus Sorghum. Utilizing germplasm homozygous for the iap allele, hybrids were readily produced between S. bicolor (2n = 20; AAB1B1) and S. macrospermum (2n = 40; WWXXYYZZ). These hybrids were intermediate to the parents in chromosome number (2n = 30) and overall morphology. Meiosis in both parents was regular; S. bicolor had 10 bivalents per pollen mother cell (PMC) and S. macrospermum had an average of 19.96 bivalents per PMC. Six hybrids were studied cytologically and meiosis was irregular, with the chromosomes associating primarily as univalents and bivalents. There was an average of 3.54 bivalents per PMC, with a range of 0-8 bivalents, most of which were rods (98%). Using FISH (fluorescent in situ hybridization), moderate levels (2.6 II per PMC) of allosyndetic recombination were observed. Genomic relationships were sufficient to assign S. macrospermum the genomic formula AAB1B1YYZZ (Y and Z remain unknown). Allosyndetic recombination in the interspecific hybrids indicates that introgression through genetic recombination should be possible if viable backcrosses can be recovered.

Apomixis, asexual reproduction through seed, is widespread among angiosperm families. Gametophytic apomixis in Pennisetum squamulatum and Cenchrus ciliaris is controlled by the apospory-specific genomic region (ASGR), which is highly conserved and macrosyntenic between these species. Thirty-two ASGR bacterial artificial chromosomes (BACs) isolated from both species and one ASGR-recombining BAC from P. squamulatum, which together cover approximately 2.7 Mb of DNA, were used to investigate the genomic structure of this region. Phrap assembly of 4,521 high-quality reads generated 1,341 contiguous sequences (contigs; 730 from the ASGR and 30 from the ASGR-recombining BAC in P. squamulatum, plus 580 from the C. ciliaris ASGR). Contigs containing putative protein-coding regions unrelated to transposable elements were identified based on protein similarity after Basic Local Alignment Search Tool X analysis. These putative coding regions were further analyzed in silico with reference to the rice (Oryza sativa) and sorghum (Sorghum bicolor) genomes using the resources at Gramene (www.gramene.org) and Phytozome (www.phytozome.net) and by hybridization against sorghum BAC filters. The ASGR sequences reveal that the ASGR (1) contains both gene-rich and gene-poor segments, (2) contains several genes that may play a role in apomictic development, (3) has many classes of transposable elements, and (4) does not exhibit large-scale synteny with either rice or sorghum genomes but does contain multiple regions of microsynteny with these species.

Crop yields are significantly reduced by aluminum toxicity on highly acidic soils, which comprise up to 50% of the world's arable land. Candidate aluminum tolerance proteins include organic acid efflux transporters, with the organic acids forming non-toxic complexes with rhizosphere aluminum. In this study, we used positional cloning to identify the gene encoding a member of the multidrug and toxic compound extrusion (MATE) family, an aluminum-activated citrate transporter, as responsible for the major sorghum (Sorghum bicolor) aluminum tolerance locus, Alt(SB). Polymorphisms in regulatory regions of Alt(SB) are likely to contribute to large allelic effects, acting to increase Alt(SB) expression in the root apex of tolerant genotypes. Furthermore, aluminum-inducible Alt(SB) expression is associated with induction of aluminum tolerance via enhanced root citrate exudation. These findings will allow us to identify superior Alt(SB) haplotypes that can be incorporated via molecular breeding and biotechnology into acid soil breeding programs, thus helping to increase crop yields in developing countries where acidic soils predominate.

Milo disease in sorghum is caused by isolates of the soil-borne fungus Periconia circinata that produce PC-toxin. Susceptibility to milo disease is conditioned by a single, semi-dominant gene, termed Pc. The susceptible allele (Pc) converts to a resistant form (pc) spontaneously at a gametic frequency of 10(-3) to 10(-4). A high-density genetic map was constructed around the Pc locus using DNA markers, allowing the Pc gene to be delimited to a 0.9 cM region on the short arm of sorghum chromosome 9. Physically, the Pc-region was covered by a single BAC clone. Sequence analysis of this BAC revealed twelve gene candidates. Several of the predicted genes in the region are homologous to disease resistance loci, including one NBS-LRR resistance gene analogue that is present in multiple tandem copies. Analysis of pc isolines derived from Pc/Pc sorghum suggests that one or more members of this NBS-LRR gene family are the Pc genes that condition susceptibility.

Genetic variation for aluminum (Al) tolerance in plants has allowed the development of cultivars that are high yielding on acidic, Al toxic soils. However, knowledge of intraspecific variation for Al tolerance control is needed in order to assess the potential for further Al tolerance improvement. Here we focused on the major sorghum Al tolerance gene, Alt ( SB ), from the highly Al tolerant standard SC283 to investigate the range of genetic diversity for Al tolerance control in sorghum accessions from diverse origins. Two tightly linked STS markers flanking Alt ( SB ) were used to study the role of this locus in the segregation for Al tolerance in mapping populations derived from different sources of Al tolerance crossed with a common Al sensitive tester, BR012, as well as to isolate the allelic effects of Alt ( SB ) in near-isogenic lines. The results indicated the existence not only of multiple alleles at the Alt ( SB ) locus, which conditioned a wide range of tolerance levels, but also of novel sorghum Al tolerance genes. Transgressive segregation was observed in a highly Al tolerant breeding line, indicating that potential exists to exploit the additive or codominant effects of distinct Al tolerance loci. A global, SSR-based, genetic diversity analysis using a broader sorghum set revealed the presence of both multiple Alt ( SB ) alleles and different Al tolerance genes within highly related accessions. This suggests that efforts toward broadening the genetic basis for Al tolerance in sorghum may benefit from a detailed analysis of Al tolerance gene diversity within subgroups across a target population.

Background: Molecular markers serve three important functions in physical map assembly. First, they provide anchor points to genetic maps facilitating functional genomic studies. Second, they reduce the overlap required for BAC contig assembly from 80 to 50 percent. Finally, they validate assemblies based solely on BAC fingerprints. We employed a six-dimensional BAC pooling strategy in combination with a high-throughput PCR-based screening method to anchor the maize genetic and physical maps., Results: A total of 110,592 maize BAC clones (approximately 6x haploid genome equivalents) were pooled into six different matrices, each containing 48 pools of BAC DNA. The quality of the BAC DNA pools and their utility for identifying BACs containing target genomic sequences was tested using 254 PCR-based STS markers. Five types of PCR-based STS markers were screened to assess potential uses for the BAC pools. An average of 4.68 BAC clones were identified per marker analyzed. These results were integrated with BAC fingerprint data generated by the Arizona Genomics Institute (AGI) and the Arizona Genomics Computational Laboratory (AGCoL) to assemble the BAC contigs using the FingerPrinted Contigs (FPC) software and contribute to the construction and anchoring of the physical map. A total of 234 markers (92.5%) anchored BAC contigs to their genetic map positions. The results can be viewed on the integrated map of maize 12., Conclusion: This BAC pooling strategy is a rapid, cost effective method for genome assembly and anchoring. The requirement for six replicate positive amplifications makes this a robust method for use in large genomes with high amounts of repetitive DNA such as maize. This strategy can be used to physically map duplicate loci, provide order information for loci in a small genetic interval or with no genetic recombination, and loci with conflicting hybridization-based information.

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

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

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

With an aim to clone the sorghum fertility restorer gene Rf1, a high-resolution genetic and physical map of the locus was constructed. The Rf1 locus was resolved to a 32-kb region spanning four open reading frames: a plasma membrane Ca(2+)-ATPase, a cyclin D-1, an unknown protein, and a pentatricopeptide repeat (PPR13) gene family member. An approximately 19-kb region spanning the cyclin D-1 and unknown protein genes was completely conserved between sterile and fertile plants as was the sequence spanning the coding region of the Ca(2+)-ATPase. In contrast, 19 sequence polymorphisms were located in an approximately 7-kb region spanning PPR13, and all markers cosegregated with the fertility restoration phenotype. PPR13 was predicted to encode a mitochondrial-targeted protein containing a single exon with 14 PPR repeats, and the protein is classified as an E-type PPR subfamily member. To permit sequence-based comparison of the sorghum and rice genomes in the Rf1 region, 0.53 Mb of sorghum chromosome 8 was sequenced and compared to the colinear region of rice chromosome 12. Genome comparison revealed a mosaic pattern of colinearity with an approximately 275-kb gene-poor region with little gene conservation and an adjacent, approximately 245-kb gene-rice region that is more highly conserved between rice and sorghum. Despite being located in a region of high gene conservation, sorghum PPR13 was not located in a colinear position on rice chromosome 12. The present results suggest that sorghum PPR13 represents a potential candidate for the sorghum Rf1 gene, and its presence in the sorghum genome indicates a single gene transposition event subsequent to the divergence of rice and sorghum ancestors.

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.

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

Linkage group identities and homologies were determined for metaphase chromosomes of Sorghum bicolor (2n = 20) by FISH of landed BACs. Relative lengths of chromosomes in FISH-karyotyped metaphase spreads of the elite inbred BTx623 were used to estimate the molecular size of each chromosome and to establish a size-based nomenclature for sorghum chromosomes (SBI-01-SBI-10) and linkage groups (LG-01 to LG-10). Lengths of arms were determined to orient linkage groups relative to a standard karyotypic layout (short arms at top). The size-based nomenclature for BTx623 represents a reasonable choice as the standard for a unified chromosome nomenclature for use by the sorghum research community.