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Results for “spatial sequencing”

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PubMed1997

Modeling of in vivo proteolytic degradation of hemoglobin.

Based on the amino acid sequences of endogenous peptides and X-ray spatial structure, mechanism of the in vivo proteolly degradation of bovine hemoglobin was analysed. The degradation was shown to be a multi-stage process. Its first stage is determined by the spatial organization of the native protein substrate, and the next stages-by the distribution of the electrostatic field potential of the protein fragments formed at the earlier stage.

Amino Acid Sequence↗

PubMed1991

Effect of differential saturation on the spatial localization performance of depth pulses.

Computer simulations of Depth pulse B (theta; (2 theta [+/- x, +/- y])2; acquire) and other Depth pulses, verified by experimental surface coil NMR studies utilizing phantom samples, reveal that the spatial localization performance of Depth pulses degrades when the repetition time is short relative to T1 because of differential saturation, i.e. T1 discrimination effects. Simulations of Depth pulse A (theta; 2 theta [+/- x, +/- y]; acquire) and Depth pulse B indicate that there is no phase-cycled pulse sequence delivery order which negates the untoward effect of T1 discrimination on spatial localization performance. The results of this study demonstrate the importance of consistent magnetization preparation prior to the delivery of each phase-cycled multiple pulse sequence in a Depth pulse cycle for obtaining optimal spatial localization performance. The untoward effects of inconsistent magnetization preparation, resulting from T1 discrimination, may be ameliorated by the application of many Depth pulse cycles.

Computer Simulation↗

PubMed2004

Comparison of the spatial limits on direction selectivity in visual areas MT and V1.

We recorded responses to apparent motion from directionally selective neurons in primary visual cortex (V1) of anesthetized monkeys and middle temporal area (MT) of awake monkeys. Apparent motion consisted of multiple stationary stimulus flashes presented in sequence, characterized by their temporal separation (delta t) and spatial separation (delta x). Stimuli were 8 degrees square patterns of 100% correlated random dots that moved at apparent speeds of 16 or 32 degrees/s. For both V1 and MT, the difference between the response to the preferred and null directions declined with increasing flash separation. For each neuron, we estimated the maximum flash separation for which directionally selective responses were observed. For the range of speeds we used, delta x provided a better description of the limitation on directional responses than did delta t. When comparing MT and V1 neurons of similar preferred speed, there was no difference in the maximum delta x between our samples from the two areas. In both V1 and MT, the great majority of neurons had maximal values of delta x in the 0.25-1 degrees range. Mean values were almost identical between the two areas. For most neurons, larger flash separations led to both weaker responses to the preferred direction and increased responses to the opposite direction. The former mechanism was slightly more dominant in MT and the latter slightly more dominant in V1. We conclude that V1 and MT neurons lose direction selectivity for similar values of delta x, supporting the hypothesis that basic direction selectivity in MT is inherited from V1, at least over the range of stimulus speeds represented by both areas.

Action Potentials↗

PubMed2005

Optical flow estimation using temporally oversampled video.

Recent advances in imaging sensor technology make high frame-rate video capture practical. As demonstrated in previous work, this capability can be used to enhance the performance of many image and video processing applications. The idea is to use the high frame-rate capability to temporally oversample the scene and, thus, to obtain more accurate information about scene motion and illumination. This information is then used to improve the performance of image and standard frame-rate video applications. This paper investigates the use of temporal oversampling to improve the accuracy of optical flow estimation (OFE). A method for obtaining high accuracy optical flow estimates at a conventional standard frame rate, e.g., 30 frames/s, by first capturing and processing a high frame-rate version of the video is presented. The method uses the Lucas-Kanade algorithm to obtain optical flow estimates at a high frame rate, which are then accumulated and refined to estimate the optical flow at the desired standard frame rate. The method demonstrates significant improvements in OFE accuracy both on synthetically generated video sequences and on a real video sequence captured using an experimental high-speed imaging system. It is then shown that a key benefit of using temporal oversampling to estimate optical flow is the reduction in motion aliasing. Using sinusoidal input sequences, the reduction in motion aliasing is identified and the desired minimum sampling rate as a function of the velocity and spatial bandwidth of the scene is determined. Using both synthetic and real video sequences, it is shown that temporal oversampling improves OFE accuracy by reducing motion aliasing not only for areas with large displacements but also for areas with small displacements and high spatial frequencies. The use of other OFE algorithms with temporally oversampled video is then discussed. In particular, the Haussecker algorithm is extended to work with high frame-rate sequences. This extension demonstrates yet another important benefit of temporal oversampling, which is improving OFE accuracy when brightness varies with time.

Algorithms↗

PubMed2000

Molecular characterization of cDNAs encoding G protein alpha and beta subunits and study of their temporal and spatial expression patterns in Nicotiana plumbaginifolia Viv.

We have isolated cDNA sequences encoding alpha and beta subunits of potential G proteins from a cDNA library prepared from somatic embryos of Nicotiana plumbaginifolia Viv. at early developmental stages. The predicted NPGPA1 and NPGPB1 gene products are 75-98% identical to the known respective plant alpha and beta subunits. Southern hybridizations indicate that NPGPA1 is probably a single-copy gene, whereas at least two copies of NPGPB1 exist in the N. plumbaginifolia genome. Northern analyses reveal that both NPGPA1 and NPGPB1 mRNA are expressed in all embryogenic stages and plant tissues examined and their expression is obviously regulated by the plant hormone auxin. Immunohistological localization of NPGPalpha1 and NPGPbeta1 preferentially on plasma and endoplasmic reticulum membranes and their immunochemical detection exclusively in microsomal cell fractions implicate membrane association of both proteins. The temporal and spatial expression patterns of NPGPA1 and NPGPB1 show conformity as well as differences. This could account for not only cooperative, but also individual activities of both subunits during embryogenesis and plant development.

Amino Acid Sequence↗

PubMed2003

Sequences within the gag gene of feline immunodeficiency virus (FIV) are important for efficient RNA encapsidation.

Feline immunodeficiency virus (FIV)-based retroviral vector systems are being developed for human gene therapy. Consequently, it has become important to know the precise sequence requirements for the packaging of FIV genome so that such sequences can be eliminated from transfer vectors post-transduction for improved safety. Recently, we have shown that sequences both within the 5'-untranslated leader region (UTR) and the 5'-end of gag are required for efficient packaging and transduction of FIV-based vectors. However, the extent of gag sequence important in the encapsidation process is not clear as well as their relative contribution to packaging. In this study, using a biologically relevant packaging system, we demonstrate that at the most 100 bp of gag sequences are sufficient for efficient RNA packaging in conjunction with the 5'-UTR and no other sequences within the next 600 bp of gag exist that affect packaging. In addition, we show that sequences within gag do not simply act as spatial elements to stabilize other structural determinants of packaging located within the 5'-UTR, but are important in themselves for the encapsidation process.

5' Untranslated Regions↗

PubMed2002

Exploring ligand-DNA space using type IIS restriction enzymes.

Investigating ligand-DNA interactions using type IIS restriction enzymes (IISRE) as footprinting reagents is reviewed and contemplated. Ligand binding at a IISRE's cleavage but not sequence recognition site protects DNA from strand scission. This spatial arrangement has been exploited in the development of qualitative (combinatorial) and quantitative ligand-DNA investigative methods collectively termed Type IIS Restriction Enzyme Footprinting (cIISREF and qIISREF respectively). In cIISREF, the consensus binding sequence of a ligand is sought by using a IISRE to segregate combinatorial library members that are bound by ligand from those that are not. A PCR is performed following the segregation step to enrich the library in ligand binding (i.e. uncut) sequences. It might be possible that diversities approaching 10(30) unique sequences could be simultaneously searched using this homogeneous and biologically relevant method. For qIISREF, a ligand-DNA equilibrium constant is measured by quantifying the amounts of target and control DNA IISRE cleavage products as a function of ligand concentration. The control sequence is engineered to not bind ligand. Along with illustrating these methods by reviewing published works, current concerns and future prospects for IISREF are discussed.

Base Sequence↗

PubMedDate not supplied

Localized proton NMR spectroscopy of brain tumors using short-echo time STEAM sequences.

Recent progress in localized proton NMR spectroscopy has been utilized to improve the spatial resolution and the metabolic specificity in a study of 19 patients with intracranial tumors. Selected examples demonstrate that short echo time stimulated echo acquisition mode sequences are able (a) to account for macroscopic tissue heterogeneity by reducing the volume of interest to 2-8 ml and (b) to facilitate a reasonable characterization of tumor metabolism by increasing the number of accessible metabolites. Proton NMR spectra were acquired within measuring times of 6.5 min on a 2.0 T whole-body system using the imaging headcoil.

Adult↗

PubMed2004

Spatial heterogeneity of crenarchaeal assemblages within mesophilic soil ecosystems as revealed by PCR-single-stranded conformation polymorphism profiling.

Microbial ecologists have discovered novel rRNA genes (rDNA) in mesophilic soil habitats worldwide, including sequences that affiliate phylogenetically within the division Crenarchaeota (domain Archaea). To characterize the spatial distribution of crenarchaeal assemblages in mesophilic soil habitats, we profiled amplified crenarchaeal 16S rDNA sequences from diverse soil ecosystems by using PCR-single-stranded-conformation polymorphism (PCR-SSCP) analysis. PCR-SSCP profiles provide a measure of relative microbial diversity in terms of richness (number of different phylotypes as estimated from the number of unique PCR-SSCP peaks) and evenness (abundance of each phylotype as estimated from the relative area under a peak). Crenarchaeal assemblages inhabiting prairie, forest, turf, and agricultural soils were characterized at six sampling locations in southern and central Wisconsin. Phylotype richness was found to be more stable than evenness among triplicate samples collected within 30 cm at each sampling location. Transformation of the PCR-SSCP data by principal-component analysis, followed by statistical testing (analysis of variance [P < 0.0001] and least-significant-difference analysis [alpha = 0.5]), supported the conclusion that each location exhibited a unique profile. To further characterize the spatial distribution of crenarchaeal assemblages at one location, additional soil samples (a total of 30) were collected from agricultural field plots at the Hancock Agricultural Research Station. PCR-SSCP revealed a patchy spatial distribution of crenarchaeal assemblages within and between these plots. This mosaic of crenarchaeal assemblages was characterized by differences in phylotype evenness that could not be correlated with horizontal distance (15 to 30 m) or with depth (0 to 20 cm below the surface). Crenarchaeal 16S rDNA clone libraries were produced and screened for unique SSCP peaks. Clones representing the dominant phylotypes at each location were identified, sequenced, and found to group phylogenetically with sequences in crenarchaeal clade C1b.

Base Sequence↗

PubMed2002

Murine peripherin gene sequences direct Cre recombinase expression to peripheral neurons in transgenic mice.

Spatially and temporally regulated somatic mutations can be achieved by using the Cre/loxP recombination system of bacteriophage P1. To develop a cell type-specific system of gene targeting in the peripheral nervous system, we generated the transgenic mouse lines expressing Cre recombinase under the control of the mouse peripherin gene promoter. The activity of the Cre recombinase during embryonic development was examined by mating the peripherin-Cre transgenic mice to the knock-in Cre-mediated recombination reporter strain, R26R. Analysis of F1 embryos from this cross showed specific excision of loxP-flanked sequences in the dorsal root ganglia, trigeminal ganglia, and olfactory epithelium, in a pattern very similar to the expression of the endogenous mouse peripherin gene, and the previously reported peripherin-lacZ transgenic mice. Thus, the peripherin-Cre mouse described here will provide a valuable tool for Cre-loxP-mediated conditional expression in the peripheral nervous system.

Animals↗

PubMed1995

Stereochemical analysis of the antigenic tip of the V3 loop peptide of HIV-1 gp120.

A novel multiple turn conformation has been observed for a segment GPGRAFY in the crystal structure of a complex of HIV-1 gp120 V3 loop peptide with the Fab fragment of a neutralizing antibody [Ghiara et al. (1994) Science 264, 82-85]. A structural motif has been defined for the peptide segment, employing idealized backbone conformations characterized by ranges of virtual C alpha torsion angles and bond angles. A search of 122 high-resolution protein crystal structures has permitted identification of 24 examples of similar structural motifs. Two major conformational families have been identified, which differ primarily in the conformation at residue 3. The observed conformation at residue 3 in family 1 is left-handed helical (alpha L) and that in family 2 is right-handed helical (alpha R). Of the 10 examples in family 1, 9 examples have Gly residues at position 3. Of the 12 examples in family 2, 7 examples have Asn/Asp at position 3. Computer modeling of the V3 loop tip sequence using the two backbone conformational families as starting points leads to minimum-energy conformations in which antigenically important side-chains occupy similar spatial arrangements. This stereochemical analysis of the V3 loop tip sequence suggests a rational basis for the design of synthetic analog peptides for use as viral antagonists or synthetic antigens.

Amino Acid Sequence↗

PubMed2006

Activation mapping as a percentage of local excitation: fMRI stability within scans, between scans and across field strengths.

Functional magnetic resonance imaging (fMRI) does not typically yield highly reproducible maps of brain activation. Maps can vary significantly even with constant scanning parameters and consistent task performance conditions (Liu et al., Magn. Reson. Med., 2004, 52:751-760). Reproducibility is even more of a problem when comparing fMRI signal magnitude and spatial extent of activation across scans involving different task performance levels, scan durations, pulse sequences or magnetic field strengths. In this report, the consistency of fMRI was reexamined by considering the relative spatial and temporal distribution of fMRI blood oxygen level dependent (BOLD) activation signals separately from the absolute magnitude of the activation signal in each brain area. Subjects repeatedly performed the same simple motor task but under a variety of imaging conditions, using both spiral and standard echo-planar pulse sequences and at 1.5- and 4.0-T magnetic field strengths. The results demonstrate that the absolute amplitude of BOLD statistical activation signals varied significantly across time and scanning conditions, but the relative spatial pattern of BOLD activation was highly reproducible across all conditions. Analysis of realistic simulated fMRI data sets indicates that stability of relative activation patterns could provide a useful tool for assessing the accuracy of fMRI maps.

Adult↗

PubMed1998

Tuning of orientation detectors in human vision.

A new method to study the tuning of orientation detectors in the human visual system is proposed. The stimulus consists of a sequence of flashed sinusoidal gratings of random orientations and spatial phases shown at a fast presentation rate. The subject's task is to report, as fast as possible, when the presence of a particular orientation (horizontal, vertical, or oblique) is seen in the stimulus sequence by pressing a button. The data are analyzed by calculating the empirical distribution of orientations present in the stimulus sequence within an optimal time-window before the button was pressed. The resulting orientation distributions show a "Mexican hat" shape, which resembles the distributions obtained in some single neurons of monkey primary visual cortex using a similar method (Ringach et al., 1997). The findings are consistent with the idea of "lateral inhibition" between neighboring detectors in the orientation domain.

Female↗

PubMed2006

Space and the parietal cortex.

Current views of the parietal cortex have difficulty accommodating the human inferior parietal lobe (IPL) within a simple dorsal versus ventral stream dichotomy. In humans, lesions of the right IPL often lead to syndromes such as hemispatial neglect that are seemingly in accord with the proposal that this region has a crucial role in spatial processing. However, recent imaging and lesion studies have revealed that inferior parietal regions have non-spatial functions, such as in sustaining attention, detecting salient events embedded in a sequence of events and controlling attention over time. Here, we review these findings and show that spatial processes and the visual guidance of action are only part of the repertoire of parietal functions. Although sub-regions in the human superior parietal lobe and intraparietal sulcus contribute to vision-for-action and spatial functions, more inferior parietal regions have distinctly non-spatial attributes that are neither conventionally 'dorsal' nor conventionally 'ventral' in nature.

Attention↗

PubMed1997

A glimpse into crabworld.

Almost all known arthropod compound eyes exhibit regional variations of resolving power, absolute light, spectral and polarisation sensitivity which are likely to be matched to the probability of significant events and the availability of cues in the visual world. To understand the signal processing requirements that have led to the evolution of matched sensory and neural filters, we thus need a detailed description of the input signals to a visual system and of the tasks to be performed under natural operating conditions. We report here on the first steps we took in an attempt to reconstruct an animal's specific visual world with emphasis on the motion domain. Fiddler crabs (genus Uca) live in burrows on sand- and mudflats and are active during low tide. They carry their eyes on long, vertically oriented stalks and use vision to detect predators and conspecific signals generated by males waving one massively enlarged claw. The crabs sit on the ground plane of a flat world, where significant events are most likely to occur in a narrow band around the horizon. We recorded scenes in a crab colony with a video camera at crab eye height. The salience of relevant features in the spatial, spectral and polarisation domains was analysed in digitised video images and short sequences of film were processed by a two-dimensional network of motion detectors at various spatial scales. The output of the network provides us with histograms of the direction and strength of motion signals in various spatio-temporal frequency bands. We discuss our results in terms of detection problems, predictability of events, global vs local information content and higher level motion processing involved in intraspecific communication.

Animals↗

PubMed2001

Adaptive vessel tracking: automated computation of vessel trajectories for improved efficiency in 2D coronary MR angiography.

A new method was investigated for improving the efficiency of ECG-gated coronary magnetic resonance angiography (CMRA) by accurate, automated tracking of the vessel motion over the cardiac cycle. Vessel tracking was implemented on a spiral gradient-echo pulse sequence with sub-millimeter in-plane spatial resolution as well as high image signal to noise ratio. Breath hold 2D CMRA was performed in 18 healthy adult subjects (mean age 46 +/- 14 years). Imaging efficiency, defined as the percentage of the slices where more than 30 mm of the vessel is visualized, was computed in multi-slice spiral scans with and without vessel tracking. There was a significant improvement in the efficiency of the vessel tracking sequence compared to the multi-slice sequence (56% vs. 32%, P < 0.001). The imaging efficiency increased further when the true motion of the coronary arteries (determined using a cross correlation algorithm) was used for vessel tracking as opposed to a linear model for motion (71% vs. 57%, P < 0.05). The motion of the coronary arteries was generally found to be linear during the systolic phase and nonlinear during the diastolic phase. The use of subject-tailored, automated tracking of vessel positions resulted in improved efficiency of coronary artery illustration on breath held 2D CMRA.

Coronary Angiography↗

PubMed2006

Development and application of three-tiered nuclear genetic markers for basal Hexapods using single-stranded conformation polymorphism coupled with targeted DNA sequencing.

BACKGROUND: Molecular genetic approaches have much to offer population biology. Despite recent advances, convenient techniques to develop and screen highly-resolving markers can be limiting for some applications and taxa. We describe an improved PCR-based, cloning-free, nuclear marker development procedure, in which single-stranded conformation polymorphism (SSCP) plays a central role. Sequence-variable alleles at putative nuclear loci are simultaneously identified and isolated from diploid tissues. Based on a multiple allele alignment, locus-specific primers are designed in conserved regions, minimizing 'null' alleles. Using two undescribed endemic Australian Collembola as exemplars, we outline a comprehensive approach to generating and validating suites of codominant, sequence-yielding nuclear loci for previously unstudied invertebrates. RESULTS: Six markers per species were developed without any baseline genetic information. After evaluating the characteristics of each new locus via SSCP pre-screening, population samples were genotyped on the basis of either DNA sequence, restriction site, or insertion/deletion variation, depending on which assay was deemed most appropriate. Polymorphism was generally high (mean of nine alleles per locus), and the markers were capable of resolving population structuring over very fine spatial scales (<100 km). SSCP coupled with targeted DNA sequencing was used to obtain genotypic, genic and genealogical information from six loci (three per species). Phylogeographic analysis identified introns as being most informative. CONCLUSION: The comprehensive approach presented here feasibly overcomes technical hurdles of (i) developing suitably polymorphic nuclear loci for non-model organisms, (ii) physically isolating nuclear allele haplotypes from diploid tissues without cloning, and (iii) genotyping population samples on the basis of nuclear DNA sequence variation.

Animals↗

PubMed1998

A specific conformation of the territory of chromosome 17 locates ERBB-2 sequences to a DNase-hypersensitive domain at the nuclear periphery.

Chromatin in interphase nuclei exhibits a topology that is associated with the transcriptional state of cells. We examined the spatial, intranuclear distribution of chromosome 17 and the ERBB-2 (HER2/neu) sequence thereon, relative to that of DNase-hypersensitive chromatin (DHC), in breast tumour cells exhibiting different levels of expression of ERBB-2. These sequences were specifically associated with the nuclear periphery, within a band of DHC. The remainder of the chromosome 17 mass showed no preferential position within the nucleus. The peripheral placement of ERBB-2 sequences is associated with a specific conformation of chromosome 17. We propose that the conformational organization of chromosome territories might represent a fundamental control mechanism in gene expression.

Breast Neoplasms↗