A reference genome for Nicotiana tabacum enables map-based cloning of homeologous loci implicated in nitrogen utilization efficiency.

BACKGROUND: Tobacco (Nicotiana tabacum) is an important plant model system that has played a key role in the early development of molecular plant biology. The tobacco genome is large and its characterisation challenging because it is an allotetraploid, likely arising from hybridisation between diploid N. sylvestris and N. tomentosiformis ancestors. A draft assembly was recently published for N. tabacum, but because of the aforementioned genome complexities it was of limited utility due to a high level of fragmentation. RESULTS: Here we report an improved tobacco genome assembly, which, aided by the application of optical mapping, achieves an N50 size of 2.17 Mb and enables anchoring of 64% of the genome to pseudomolecules; a significant increase from the previous value of 19%. We use this assembly to identify two homeologous genes that explain the differentiation of the burley tobacco market class, with potential for greater understanding of Nitrogen Utilization Efficiency and Nitrogen Use Efficiency in plants; an important trait for future sustainability of agricultural production. CONCLUSIONS: Development of an improved genome assembly for N. tabacum enables what we believe to be the first successful map-based gene discovery for the species, and demonstrates the value of an improved assembly for future research in this model and commercially-important species.

Ricebase: a breeding and genetics platform for rice, integrating individual molecular markers, pedigrees and whole-genome-based data.

Ricebase (http://ricebase.org) is an integrative genomic database for rice (Oryza sativa) with an emphasis on combining datasets in a way that maintains the key links between past and current genetic studies. Ricebase includes DNA sequence data, gene annotations, nucleotide variation data and molecular marker fragment size data. Rice research has benefited from early adoption and extensive use of simple sequence repeat (SSR) markers; however, the majority of rice SSR markers were developed prior to the latest rice pseudomolecule assembly. Interpretation of new research using SNPs in the context of literature citing SSRs requires a common coordinate system. A new pipeline, using a stepwise relaxation of stringency, was used to map SSR primers onto the latest rice pseudomolecule assembly. The SSR markers and experimentally assayed amplicon sizes are presented in a relational database with a web-based front end, and are available as a track loaded in a genome browser with links connecting the browser and database. The combined capabilities of Ricebase link genetic markers, genome context, allele states across rice germplasm and potentially user curated phenotypic interpretations as a community resource for genetic discovery and breeding in rice.

Quantitative computer-assisted osteodensitometry in total hip arthroplasty.

Several factors can cause bone loss and fixation failure following total hip arthroplasty (THA), including polyethylene wear debris, implant micromotion and stress shielding. Various techniques have been used in an effort to detect bone density loss in vivo, all with varying success. Quantitative computed tomography (qCT)-assisted osteodensitometry has been shown to be useful in assessing the in vivo structural bone changes after THA. It has a high resolution, accuracy and reproducibility, thereby making it a useful tool for research purposes, and it is able to differentiate between cortical and cancellous bone structures and assess the bone/implant interface. This technique also provides valuable information about the pattern of stress shielding which occurs around the prosthesis and can show early bony changes, which may prove informative about the quality of implant fixation and surrounding bone adaptation. In conjunction with finite-element analysis, qCT is able to generate accurate patient-specific meshes on which to model implants and their effect on bone remodelling. This technology can be useful to predict bone remodelling and the quality of implant fixation using prostheses with different design and/or biomaterials. In the future, this tool could be used for pre-clinical validation of new implants before their introduction in the market-place.

The phenomenon and efficiency of ligamentotaxis after dorsal stabilization of thoracolumbar burst fractures.

Thirty-six consecutive patients with burst fractures of the thoracolumbar spine and with a fractured posterior vertebral surface dislocated into the spinal canal without neurological symptoms were treated with the AO internal fixator. Computed tomography-aided planimetry of the spinal canal was undertaken preoperatively and within 1 week postoperatively to elucidate the effect of kyphosis correction and distraction on spinal canal widening (ligamentotaxis). The stenosis of the spinal canal area (SCA) was reduced from 29% preoperatively to 19% postoperatively (+10%) of the estimated original area, and the stenosis of the mid-sagittal diameter (MSD) reduced from 31 to 23% (+8%). The widening of the SCA was greater at the level of L1/L2 (+13%) than at L3/L4 (+6%). High preoperative canal compromise was associated with greater absolute spinal canal widening. Large trapezoid-shaped fragments resisted reduction by ligamentotaxis. Even though the effect of ligamentotaxis after operative treatment with the internal fixator was proven, a certain stenosis of the spinal canal remains in most cases. Especially for patients with fracture-related neurological symptoms, ligamentotaxis alone does not seem sufficient for the requested spinal decompression. Even an exact analysis of preoperative CT scans under consideration of the fracture level will not always allow an exact prognosis of the expected effect of ligamentotaxis.

Characteristics of the tomato nuclear genome as determined by sequencing undermethylated EcoRI digested fragments.

A collection of 9,990 single-pass nuclear genomic sequences, corresponding to 5 Mb of tomato DNA, were obtained using methylation filtration (MF) strategy and reduced to 7,053 unique undermethylated genomic islands (UGIs) distributed as follows: (1) 59% non-coding sequences, (2) 28% coding sequences, (3) 12% transposons-96% of which are class I retroelements, and (4) 1% organellar sequences integrated into the nuclear genome over the past approximately 100 million years. A more detailed analysis of coding UGIs indicates that the unmethylated portion of tomato genes extends as far as 676 bp upstream and 766 bp downstream of coding regions with an average of 174 and 171 bp, respectively. Based on the analysis of the UGI copy distribution, the undermethylated portion of the tomato genome is determined to account for the majority of the unmethylated genes in the genome and is estimated to constitute 61+/-15 Mb of DNA (approximately 5% of the entire genome)--which is significantly less than the 220 Mb estimated for gene-rich euchromatic arms of the tomato genome. This result indicates that, while most genes reside in the euchromatin, a significant portion of euchromatin is methylated in the intergenic spacer regions. Implications of the results for sequencing the genome of tomato and other solanaceous species are discussed.

The Arabidopsis Information Resource (TAIR): a comprehensive database and web-based information retrieval, analysis, and visualization system for a model plant.

Arabidopsis thaliana, a small annual plant belonging to the mustard family, is the subject of study by an estimated 7000 researchers around the world. In addition to the large body of genetic, physiological and biochemical data gathered for this plant, it will be the first higher plant genome to be completely sequenced, with completion expected at the end of the year 2000. The sequencing effort has been coordinated by an international collaboration, the Arabidopsis Genome Initiative (AGI). The rationale for intensive investigation of Arabidopsis is that it is an excellent model for higher plants. In order to maximize use of the knowledge gained about this plant, there is a need for a comprehensive database and information retrieval and analysis system that will provide user-friendly access to Arabidopsis information. This paper describes the initial steps we have taken toward realizing these goals in a project called The Arabidopsis Information Resource (TAIR) (www.arabidopsis.org).

AN9, a petunia glutathione S-transferase required for anthocyanin sequestration, is a flavonoid-binding protein.

AN9 is a glutathione S-transferase from petunia (Petunia hybrida) required for efficient anthocyanin export from the site of synthesis in the cytoplasm into permanent storage in the vacuole. For many xenobiotics it is well established that a covalent glutathione (GSH) tag mediates recognition of molecules destined for vacuolar sequestration by a tonoplast-localized ATP-binding cassette pump. Here we inquired whether AN9 catalyzes the formation of GSH conjugates with flavonoid substrates. Using high-performance liquid chromatography analysis of reaction mixtures containing enzyme, GSH, and flavonoids, including anthocyanins, we could detect neither conjugates nor a decrease in the free thiol concentration. These results suggest that no conjugate is formed in vitro. However, AN9 was shown to bind flavonoids using three assays: inhibition of the glutathione S-transferase activity of AN9 toward the common substrate 1-chloro 2,4-dinitrobenzene, equilibrium dialysis, and tryptophan quenching. We conclude that AN9 is a flavonoid-binding protein, and propose that in vivo it serves as a cytoplasmic flavonoid carrier protein.

NPS 1506: a novel NMDA receptor antagonist: neuroprotective effects in a model of closed head trauma in rats.

We examined whether NPS 1506, a novel uncompetitive N-methyl-D-aspartate receptor antagonist, influences neurological outcome following closed head trauma (CHT) in rats. One hundred ten rats were divided into 11 groups: CHT (yes/no), treatment with NPS 1506 (yes/no), and time of euthanization (24 h/48 h). The dose of NPS 1506 was 1 mg/kg IV at 1 and 4 hours following CHT or sham operation. Closed head trauma induced the following changes in the injured hemisphere: Decreased specific gravity (sg) (1.036 +/- 0.006) and magnesium (Mg) (0.042 +/- 0.005 microg/mg) at 24 hours, and potassium (K) at 24 (1.145 +/- 0.376 microg/mg) and 48 hours, and increased water content (W) (84.9 +/- 2.5%) and sodium (Na) (2.135 +/- 0.699 microg/mg) at 24 hours, and calcium (Ca) at 24 (0.543 +/- 0.157 microg/mg) and 48 hours. These were reversed by NPS 1506; sg of 1.043 +/- 0.004, Mg of 0.077 +/- 0.009 microg/mg, K of 1.930 +/- 0.238 microg/mg, W of 81.5 +/- 1.9%, Ca of 0.043 +/- 0.023 microg/mg, and Na of 0.688 +/- 0.110 microg/mg. In groups not given NPS 1506, a nonsignificant decrease in neurological severity score (NSS) occurred at 24 and 48 hours as compared to NSS at 1 hour after CHT. In groups given NPS 1506, NSS at 24 and 48 hours decreased significantly (improved) compared to NSS at 1 hour, but not compared to NSS at 24 and 48 hours in groups not given NPS 1506. NPS 1506 caused no significant change in ischemic tissue volume or hemorrhagic necrosis volume in the injured hemisphere at 24 hours or 48 hours. These findings indicate that NPS 1506 improved measures of brain tissue edema (at 24 hours but not at 48 hours) and ion homeostasis, and this improvement was not related to other measures of outcome.

The cost of sealant application in a state dental disease prevention program.

A school-based pit-and-fissure sealant activity was developed by the Dental Health Section, Health Services Division, New Mexico Health and Environment Department as an adjunct to an ongoing dental disease prevention program. The sealants were applied by dental hygienist and dental assistant teams using a "four-handed" technic. The teams took portable dental equipment directly into the schools to apply the sealants to newly erupted permanent teeth. The costs to seal 15,281 teeth in 3,272 children were calculated to be $1.59 per tooth, and $7.41 per child. The average time for sealant application was nine minutes, 24.6 seconds per child. The one-year retention rate was 83 percent complete retention and 10 percent partial retention.