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Results for “spatial sequencing”

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PubMed1998

Dynamic contrast-enhanced MR imaging of the entire breast with spectral-selective inversion fast three dimensional sequence.

Dynamic contrast-enhanced images with high spatial and temporal resolutions were acquired with a fast 3D spoiled gradient echo (SPGR) sequence using spectral selective inversion recovery (IR) pulse. Five healthy volunteers and 12 patients with 14 pathologically proven breast lesions were studied. Fat suppressed volume image data covering the entire breast were obtained with a sufficient spatial resolution (0.9 x 1.5 x 3.0 mm3) and an imaging time of 57 s. By using the criteria including peripheral enhancement and presence of spiculation, sensitivity, specificity, and accuracy in detecting malignant lesions were 88.9, 80.0 and 85.7%, respectively. Although the C/N and S/N ratios were approximately 30% less than those of the conventional fat suppressed 3D technique, fast 3D SPGR imaging with spectral IR method demonstrated sufficient image quality for both time intensity analysis and morphological evaluation of the breast lesions with a data acquisition time less than half of the conventional method. This technique can substantially improve spatial and temporal resolutions of dynamic MR images of the breast and will be useful in evaluating malignant and benign breast lesions.

Adipose Tissue↗

PubMed2000

High throughput cellular localization of specific plant mRNAs by liquid-phase in situ reverse transcription-polymerase chain reaction of tissue sections.

Advances in high throughput DNA sequencing and bioinformatic gene discovery far outpace our ability to analyze gene function, necessitating development of more efficient means to examine expression at the cellular level. Here we present a polymerase chain reaction-based method to detect mRNA species in situ in which essentially all of the steps are carried out in liquid phase in a 96-well microtiter tray and only the final signal detection is performed on a microscope slide. We demonstrate the sensitivity of the method by the cellular localization of mRNA for the Tkn2 transcription factor in a wide variety of plant tissues, and its selectivity in discriminating a single gene family member by the in situ localization of rbcs3 transcripts. Furthermore, we demonstrate the utility of the in-well in situ method in detecting FDL and IFL1 transcripts in Arabidopsis sections, thus establishing the method as a tool to determine spatial expression pattern of sequences obtained from genomic sequencing projects. Being amenable to robotic processing, in-well in situ reverse transcription-polymerase chain reaction permits a great enhancement in the number of tissue samples that can be processed. Consequently, this method may become a powerful tool for functional genomics studies, permitting the cellular site of transcription of large numbers of sequences obtained from databases to be rapidly established.

Arabidopsis↗

PubMed2005

Wave-like spread of Ebola Zaire.

In the past decade the Zaire strain of Ebola virus (ZEBOV) has emerged repeatedly into human populations in central Africa and caused massive die-offs of gorillas and chimpanzees. We tested the view that emergence events are independent and caused by ZEBOV variants that have been long resident at each locality. Phylogenetic analyses place the earliest known outbreak at Yambuku, Democratic Republic of Congo, very near to the root of the ZEBOV tree, suggesting that viruses causing all other known outbreaks evolved from a Yambuku-like virus after 1976. The tendency for earlier outbreaks to be directly ancestral to later outbreaks suggests that outbreaks are epidemiologically linked and may have occurred at the front of an advancing wave. While the ladder-like phylogenetic structure could also bear the signature of positive selection, our statistical power is too weak to reach a conclusion in this regard. Distances among outbreaks indicate a spread rate of about 50 km per year that remains consistent across spatial scales. Viral evolution is clocklike, and sequences show a high level of small-scale spatial structure. Genetic similarity decays with distance at roughly the same rate at all spatial scales. Our analyses suggest that ZEBOV has recently spread across the region rather than being long persistent at each outbreak locality. Controlling the impact of Ebola on wild apes and human populations may be more feasible than previously recognized.

Animals↗

PubMed1984

Spatial or flow velocity phase encoding gradients in NMR imaging.

We describe time modulated field gradient sequences able to selectively phase encode spatial location or flow velocity in NMR Signals. Two flow conditions are presented: constant flow velocity and simple harmonic flow superposed on steady flow. In each case we show that specific modulated gradients are available in order to discriminate between stationary and mobile protons. The methods are presented in the one dimensional case. The advantages of the phase modulation for flow analysis are the following: the method is sensitive to flow direction, unaffected by the T2, and the stationary and flow parameters are quantifiable factors.

Blood Circulation↗

PubMed2006

Microtubule stability studied by three-dimensional molecular theory of solvation.

We study microtubular supramolecular architectures of tubulin dimers self-assembling into linear protofilaments, in turn forming a closed tube, which is an important component of the cytoskeleton. We identify the protofilament arrangements with the lowest free energy using molecular dynamics to optimize tubulin conformations. We then use the three-dimensional molecular theory of solvation to obtain the hydration structure of protofilaments built of optimized tubulins and the solvent-mediated effective potential between them. The latter theoretical method, based on first principles of statistical mechanics, is capable of predicting the structure and thermodynamics of solvation of supramolecular architectures. We obtained a set of profiles of the potential of mean force between protofilaments in a periodic two-dimensional sheet in aqueous solution. The profiles were calculated for a number of amino acid sequences, tubulin conformations, and spatial arrangements of protofilaments. The results indicate that the effective interaction between protofilaments in aqueous solution depends little on the isotypes studied; however, it strongly depends on the M loop conformation of beta-tubulin. Based on the analysis of the potential of mean force between adjacent protofilaments, we found the optimal arrangement of protofilaments, which is in good agreement with other studies. We also decomposed the potential of mean force into its energetic and entropic components, and found that both are considerable in the free-energy balance for the stabilized protofilament arrangements.

Computer Simulation↗

PubMed2000

Characterization of the potato MADS-box gene STMADS16 and expression analysis in tobacco transgenic plants.

A new MADS-box gene, STMADS16, has been cloned in Solanum tuberosum L. that is expressed in all vegetative tissues of the plant, mainly in the stem, but not in flower organs. STMADS16 expression is established early during vegetative development and is not regulated by light. Sequence similarity besides the spatial and temporal expression patterns allow to define a novel MADS-box subfamily comprising STMADS16 and the gene STMADS11. Expression of the STMADS16 sense cDNA under the control of the 35S cauliflower mosaic virus promoter modifies the inflorescence structure by increasing both internode length and flower proliferation of the inflorescence meristems, and confers vegetative features to the flower. Moreover, STMADS16 ectopic expression overcomes the increase in flowering time and node number produced under short-day photoperiod, while the flowering time is not affected in long-day conditions. These results are discussed in terms of a possible role for STMADS16 in promoting vegetative development.

Amino Acid Sequence↗

PubMed2006

Linking cDNA-AFLP-based gene expression patterns and ESTs.

Massive amounts of DNA sequence data, generated from expressed sequence tag (EST) and genome sequencing projects, require efficient methods to link sequence databases with temporal and spatial expression profiles. To meet this need, we have developed a powerful computer program (GenEST), which links cDNA sequence data (including EST sequences) with transcript profiles revealed by cDNA-amplified fragment length polymorphism (AFLP). cDNA-AFLP is a highly reproducible differential display method based on restriction enzyme digests and selective amplification under high stringency conditions. GenEST predicts the sizes of virtual transcript derived fragments (TDFs) from cDNA sequences digested in silico. The resulting virtual TDFs could be traced back among the thousands of TDFs displayed on cDNA-AFLP gels. As a consequence, cDNA sequence databases can be screened very efficiently to identify genes with relevant expression profiles. Vice versa, using the restriction enzyme recognition sites, the primer extensions and the estimated TDF size as identifiers, the DNA sequence(s) corresponding to a TDF with an interesting expression pattern can be identified.

Automation↗

PubMed2004

Corsi's block-tapping test: some characteristics of the spatial path which influence memory.

By using a sequence of digits it is possible to indicate the path of Corsi's block-tapping test by drawing a line to join the central points of the cubes which make up the sequence. By doing this one can analyze the sequence of digits according to quantitative parameters. There were three characteristics of the spatial path taken into consideration in this research: the number of cubes which constitute a series of digits, the number of times the path intersected itself, and the length of the path measured in millimeters. The experiment, carried out with 70 university students, showed that all three factors were significant on analysis of variance, and also that there were differences between the sexes, the men performing better. No interactions were significant. Despite this, additional significant differences were found among the series with the same number of cubes, intersections, and length, meaning that other variables influence the difficulty of the spatial path.

Adult↗

PubMed2005

BjalphaIT: a novel scorpion alpha-toxin selective for insects--unique pharmacological tool.

Long-chain neurotoxins derived from the venom of the Buthidae scorpions, which affect voltage-gated sodium channels (VGSCs) can be subdivided according to their toxicity to insects into insect-selective excitatory and depressant toxins (beta-toxins) and the alpha-like toxins which affect both mammals and insects. In the present study by the aid of reverse-phase HPLC column chromatography, RT-PCR, cloning and various toxicity assays, a new insect selective toxin designated as BjalphaIT was isolated from the venom of the Judean Black Scorpion (Buthotus judaicus), and its full primary sequence was determined: MNYLVVICFALLLMTVVESGRDAYIADNLNCAYTCGSNSYCNTECTKNGAVSGYCQWLGKYGNACWCINLPDKVPIRIPGACR (leader sequence is underlined). Despite its lack of toxicity to mammals and potent toxicity to insects, BjalphaIT reveals an amino acid sequence and an inferred spatial arrangement that is characteristic of the well-known scorpion alpha-toxins highly toxic to mammals. BjalphaITs sharp distinction between insects and mammals was also revealed by its effect on sodium conductance of two cloned neuronal VGSCs heterloguously expressed in Xenopus laevis oocytes and assayed with the two-electrode voltage-clamp technique. BjalphaIT completely inhibits the inactivation process of the insect para/tipE VGSC at a concentration of 100 nM, in contrast to the rat brain Na(v)1.2/beta1 which is resistant to the toxin. The above categorical distinction between mammal and insect VGSCs exhibited by BjalphaIT enables its employment in the clarification of the molecular basis of the animal group specificity of scorpion venom derived neurotoxic polypeptides and voltage-gated sodium channels.

Amino Acid Sequence↗

PubMed1994

Fluorescence resonance energy transfer within the regulatory light chain of myosin.

Rabbit skeletal muscle myosin regulatory light chain-2 (LC2) contains two reactive cysteine residues, Cys125 and Cys154, and one tryptophan at position 137. Using wild-type rabbit LC2 or its genetically engineered mutant with Cys125-->Arg (C125R), these residues can be selectively modified with fluorescent or chromophoric probes for spectroscopic studies. We have bound suitable donor/acceptor probe pairs to the two cysteine residues and Trp137 in LC2 or C125R, and measured the distance in solution between the probes by fluorescence resonance energy transfer spectroscopy. C125R was made to facilitate specific labelling of the less reactive Cys154, thus allowing the distance between Cys154 and Trp137 to be measured. Our measurements show that these residues are in close proximity to each other, the distance between them ranging from 1.7 nm (between Cys125 and Trp137) to 2.7 nm (Cys125 and Cys154). These results suggest that Cys125, Trp137 and Cys154, spanning up to 29 residues in the sequence of LC2, are spatially close, consistent with these residues residing within a C-terminal globular domain. The distances we obtained are in agreement with previous crosslinking studies [Huber, P. A., Brunner, U.T. & Schaub, M. C. (1989) Biochemistry 28, 9116-9123; Saraswat, L. & Lowey, S. (1991) J. Biol. Chem. 266, 19777-19785] and structure predictions of LC2. LC2 is located at the head-rod junction of the myosin crossbridge, and provides the primary regulatory mechanism in molluscan and smooth muscle. In skeletal muscle, its functional role is unclear, although it has been implicated in modulating actomyosin interaction [Metzger, J. M. & Moss, R. L. (1992) Biophys. J. 63, 460-468]. The incorporation of spectroscopic probes onto the light chains of myosin in solution or in fibres has become a valuable tool for evaluating the dynamic properties of the crossbridge during force generation.

Amino Acid Sequence↗

PubMed1992

MR imaging of the lungs: value of short TE spin-echo pulse sequences.

OBJECTIVE: An experimental short echo delay (TE = 7 msec) T1-weighted spin-echo sequence was compared with a conventional (TE = 20 msec) T1-weighted spin-echo sequence in the assessment of normal and abnormal lung parenchyma. Comparison was also made with high-resolution CT of abnormal lung parenchyma. SUBJECTS AND METHOD: At 1.5 T, an experimental short echo delay T1-weighted multislice spin-echo sequence (TR = RR interval, TE = 7 msec) was compared with an optimal conventional T1-weighted spin-echo sequence (TR = RR interval, TE = 20 msec, spatial presaturation). Ten healthy volunteers were examined with both sequences. The mean signal intensity and signal-to-noise ratios were calculated in lung parenchyma for both sequences. Two radiologists compared the visualization of normal lung parenchymal structures with the two techniques. In 24 patients with diffuse lung disease, results with both MR sequences and with high-resolution CT were compared. RESULTS: The signal intensity was significantly greater (p < .001) with the TE of 7 msec than with the TE of 20 msec, resulting in a 3.5-fold improvement in the signal-to-noise ratio. The 7-msec TE improved visualization of lung parenchymal structures, including peripheral vessels, interlobular septa or veins, and centrilobular arteries. In the patients with diffuse lung disease, pulmonary parenchymal abnormalities were better visualized on the images with TEs of 7 msec than on images with TEs of 20 msec. When compared with high-resolution CT, the sequence with a TE of 7 msec provided comparable assessment of air-space opacification and dense consolidation, but it was inferior to high-resolution CT in the anatomic assessment of lung parenchyma. CONCLUSION: This experimental spin-echo sequence with a TE of 7 msec significantly improves the signal-to-noise ratio, allowing improved visualization of normal and abnormal pulmonary parenchyma when compared with conventional spin-echo images with a TE of 20 msec. Although anatomic detail remains inferior to that seen with high-resolution CT, the improved image quality with a TE of 7 msec suggests that assessment and follow-up of parenchymal lung disease might be possible with MR, thereby avoiding ionizing radiation.

Adult↗

PubMed1998

Synthetic integrin-binding peptides promote adhesion and proliferation of human periodontal ligament cells in vitro.

Periodontal ligament (PDL) cells have been shown to express several integrins (alphav, alpha5, beta1, beta3) that use RGD (arginine-glycine-aspartic Acid)-dependent mechanisms for the recognition and binding of their ligands. The objective of this study was to evaluate the effects of certain integrin-binding cyclic and linear synthetic RGD-containing peptides on PDL cells' adhesion, proliferation, and de novo protein synthesis in vitro. Fifth passages of normal human PDL cells established from teeth extracted from patients (ages 12 to 14) for orthodontic reasons were used for all experiments. Synthetic peptides containing the EPRGDNYR sequence in two different spatial conformations (linear and cyclic) were covalently attached to bovine serum albumin (BSA). Type I collagen, EPRGDNYR-BSA conjugates, 1:1 mixtures of type I collagen and conjugates, as well as BSA (a negative control) were coated on bacteriological plastic and evaluated for their attachment-promoting activities. In addition, the effects of these substrates on cell proliferation were evaluated by [3H]thymidine incorporation by the PDL cells. For attachment and spreading, the cyclic forms of EPRGDNYR-BSA conjugate and type I collagen were most potent, followed by linear EPRGDNYR-BSA conjugate. The effects of all collagen/conjugate mixtures were equivalent to that of type I collagen except for the collagen/linear EPRGDNYR-BSA mixture, which was less potent. The cyclic EPRGDNYR-BSA conjugate was the most effective substrate to stimulate cell proliferation, and it was followed in potency by the linear peptide-BSA conjugate. Collagen alone did not stimulate [3H]thymidine incorporation above the control level. Mixtures of collagen with all of the conjugates showed stimulatory effects similar to that of the cyclic peptide-BSA conjugate. No significant differences in de novo protein synthesis were detected. These results suggest that the synthetic RGD-containing peptides attached to a carrier are potent ligands for the human PDL cells, and that they could provide a basis for the development of new strategies aimed at the regeneration of the periodontium.

Adolescent↗

PubMed1988

Prediction of the location of structural domains in globular proteins.

The location of structural domains in proteins is predicted from the amino acid sequence, based on the analysis of a computed contact map for the protein, the average distance map (ADM). Interactions between residues i and j in a protein are subdivided into several ranges, according to the separation [i-j[ in the amino acid sequence. Within each range, average spatial distances between every pair of amino acid residues are computed from a data base of known protein structures. Infrequently occurring pairs are omitted as being statistically insignificant. The average distances are used to construct a predicted ADM. The ADM is analyzed for the occurrence of regions with high densities of contacts (compact regions). Locations of rapid changes of density between various parts of the map are determined by the use of scanning plots of contact densities. These locations serve to pinpoint the distribution of compact regions. This distribution, in turn, is used to predict boundaries of domains in the protein. The technique provides an objective method for the location of domains both on a contact map derived from a known three-dimensional protein structure, the real distance map (RDM), and on an ADM. While most other published methods for the identification of domains locate them in the known three-dimensional structure of a protein, the technique presented here also permits the prediction of domains in proteins of unknown spatial structure, as the construction of the ADM for a given protein requires knowledge of only its amino acid sequence.

Adenylate Kinase↗

PubMed1984

Fast computer search for similar DNA sequences.

An extremely fast method of searching a nucleic acid sequence database against a probe sequence is described. The method is based on the detection of deviation from expected number and deviation from random spatial distribution of sub-sequences which are unique within a sequence, and shared between that sequence and the probe. On an IBM 3081 computer, total search of an encoded form of the EMBL nucleic acid sequence database with a 1 kbase probe sequence is completed in a few seconds. Previous best methods for a similar task required a few minutes.

Animals↗

PubMedDate not supplied

Use of magnetic resonance imaging in spinal trauma: indications, techniques, and utility.

Magnetic resonance (MR) imaging of acute spinal injury provides excellent visualization of neurologic and soft-tissue structures in a noninvasive format. Advances in imaging-sequence techniques have made possible more rapid acquisition of images with greater spatial resolution. Appropriate selection of imaging sequences allows improved imaging and contrast of the pathologic processes involved in acute spinal trauma, including spinal cord, soft-tissue, and ligamentous injury. Three patterns of spinal cord injury have been identified. Type I is representative of acute cord hemorrhage. Type II represents spinal cord edema. Type III is a mixed hemorrhagic-edematous presentation. Correlation of MR findings with experimental and clinical spinal cord injury has given a relative predictive value to spinal cord injury patterns on MR images indicative of long-term neurologic outcome. Magnetic resonance imaging is useful in delineating soft-tissue injuries associated with spinal column trauma. Despite the improved spatial resolution of MR imaging, plain radiography and computed tomography remain the standard modalities for visualizing spinal fractures.

Acute Disease↗

PubMed2005

[Spatial delimitation of low grade oligodendrogliomas].

Image-guided surgery is the central element of therapeutic management of low grade gliomas and consequently, a precise preoperative definition of their spatial extension is necessary. The question of the present work is: do the imaging abnormalities delineate the real spatial development of low grade oligodendrogliomas? A review of the literature showed that MRI on T2-weighted and FLAIR sequences are used to delineate the spatial developement of these tumours and that spectroscopic magnetic resonance imaging is more sensible to appreciate it. Moreover, mathematical models and histological studies suggest that MRI does not indicate the actual spatial extension of low grade oligodendrogliomas. This study focused on histological analysis of biopsy samples performed outside MRI imaging abnormalities in patients who harboured a low grade oligodendroglioma. It showed that isolated tumour cells were identified beyond imaging abnormalities in all of the 17 patients studied. In 15 of those 17 patients, isolated tumour cells were identified in the most distant biopsy samples taken outside imaging abnormalities. Thus, conventional imaging findings, including MRI on T2-weighted and FLAIR sequences, are not able to provide the real spatial development and boundaries of low grade oligodendrogliomas.

Adolescent↗

PubMed2000

Developmentally regulated expression of Thy-1 in structures of the mouse sensory-motor system.

Thy-1 is a cell-surface molecule of the immunoglobulin superfamily which is expressed at high levels in the mature nervous system. Thy-1 has been implicated in regulating axonal outgrowth and synaptic function, but little is known regarding its cellular localization and expression in the central nervous system (CNS) during development or in adulthood. In this study, Thy-1 gene expression and protein localization were examined in sensory-motor and related areas of the adult and postnatally developing mouse CNS. Thy-1 mRNA expression was restricted to neurons; immunoreactivity was densely distributed throughout the neuropil of all regions examined, often delineated the neuronal plasmalemma, and labeled axons in white matter tracts of the brain and spinal cord. In adulthood, immunolabeling was regionally widespread and was present relatively homogeneously throughout all cell-dense layers of sensory-motor cortex, throughout most thalamic nuclei, globus pallidus, and spinal cord. Developmentally, however, Thy-1 expression and localization exhibited a spatially and temporally staggered sequence leading to the adult pattern. In sensory-motor cortex, Thy-1 expression in layer V preceded expression in other layers; in the barrel field, labeling of barrel septa preceeded a gradually increasing intensity of immunolabeling of barrel centers; in the thalamus, Thy-1 exhibited a differential onset and temporal pattern of expression across different nuclei associated with motor, sensory, or limbic systems; in the caudate nucleus, Thy-1 expression was greatest during the first postnatal week of life before declining during subsequent development. Taken together, the adult distribution and developmental patterns leading to it form a unique profile in comparison with other structurally related glycosyl-phosphatidylinositol (GPI)-anchored neural cell adhesion molecules. The pattern and timing of Thy-1 expression across layers and nuclei during early postnatal development are more complex than previously recognized, thus perhaps reflecting varied roles for Thy-1 in aspects of structural or functional maturation which proceed independently of the timing of neurogenesis, migration, and dendritic and axonal growth.

Animals↗

PubMed2000

A new fast and unsynchronized method for MRI of viscoelastic properties of soft tissues.

Quantitative measurement of mechanical properties of biologic tissues may have several applications for diagnosis or biomechanic modeling in sports medicine, traumatology, or computer-guided surgery. The magnetic resonance imaging (MRI) methods previously tested for these applications all required synchronization between MRI acquisition pulses and the mechanical stimulation. A new unsynchronized method operating with no prior knowledge of intensity, direction, and frequency of the mechanical waves is proposed. A specifically modified SPAMM (SPAtial Modulation of Magnetization) sequence has been used, operating on a 0.2-T MRI system. The experimental results obtained on test objects fit well with theoretical calculations. The new proposed method is very fast (a less than 5-second acquisition time) for routine clinical use.

Biomechanical Phenomena↗