Gene enrichment using antibodies to DNA/RNA hybrids : mapping the ribosomal DNA of slime mold and rat

A novel method for gene enrichment has been developed and applied to mapping the rRNA genes of two eucaryotic organisms. The method makes use of antibodies to DNA/RNA hybrids prepared by injecting rabbits with the synthetic hybrid poly(rA)•poly(dT). Antibodies which cross-react with non-hybrid nucleic acids were removed from the purified IgG fraction by adsorption on columns of DNA-Sepharose, oligo(dT)-cellulose, and poly(rA)-Sepharose. Subsequent purification of the specific DNA/RNA hybrid antibody was carried out on a column of oligo(dT)-cellulose to which poly(rA) was hybridized. Attachment of these antibodies to CNBr-activated Sepharose produced an affinity resin which specifically binds DNA/RNA hybrids.

In order to map the rDNA of the slime mold Dictyostelium discoideum, R-loops were formed using unsheared nuclear DNA and the 178 and 268 rRNAs of this organism. This mixture was passed through a column containing the affinity resin, and bound molecules containing R- loops were eluted by high salt. This purified rDN A was observed directly in the electron microscope. Evidence was obtained that there is a physical end to Dictyostelium rDN A molecules approximately 10 kilobase pairs (kbp) from the region which codes for the 268 rRNA. This finding is consistent with reports of other investigators that the rRNA genes exist as inverse repeats on extra-chromosomal molecules of DNA unattached to the remainder of the nuclear DNA in this organism.

The same general procedure was used to map the rRNA genes of the rat. Molecules of DNA which contained R-loops formed with the 188 and 288 rRNAs were enriched approximately 150- fold from total genomal rat DNA by two cycles of purification on the affinity column. Electron microscopic measurements of these molecules enabled the construction of an R-loop map of rat rDNA. Eleven of the observed molecules contained three or four R-loops or else two R-loops separated by a long spacer. These observations indicated that the rat rRNA genes are arranged as tandem repeats. The mean length of the repeating units was 37.2 kbp with a standard deviation of 1.3 kbp. These eleven molecules may represent repeating units of exactly the same length within the errors of the measurements, although a certain degree of length heterogeneity cannot be ruled out. If significantly shorter or longer repeating units exist, they are probably much less common than the 37.2 kbp unit.

The last section of the thesis describes the production of antibodies to non-histone chromosomal proteins which have been exposed to the ionic detergent sodium dodecyl sulfate (SDS). The presence of low concentrations of SDS did not seem to affect either production of antibodies or their general specificity. Also, a technique is described for the in situ immunofluorescent detection of protein antigens in polyacrylamide gels.

Part I. A study of the velocity structure of the earth by the use of core phases. Part II. The 1971 San Fernando earthquake series focal mechanisms and tectonics

The initial objective of Part I was to determine the nature of upper mantle discontinuities, the average velocities through the mantle, and differences between mantle structure under continents and oceans by the use of P'dP', the seismic core phase P'P' (PKPPKP) that reflects at depth d in the mantle. In order to accomplish this, it was found necessary to also investigate core phases themselves and their inferences on core structure. P'dP' at both single stations and at the LASA array in Montana indicates that the following zones are candidates for discontinuities with varying degrees of confidence: 800-950 km, weak; 630-670 km, strongest; 500-600 km, strong but interpretation in doubt; 350-415 km, fair; 280-300 km, strong, varying in depth; 100-200 km, strong, varying in depth, may be the bottom of the low-velocity zone. It is estimated that a single station cannot easily discriminate between asymmetric P'P' and P'dP' for lead times of about 30 sec from the main P'P' phase, but the LASA array reduces this uncertainty range to less than 10 sec. The problems of scatter of P'P' main-phase times, mainly due to asymmetric P'P', incorrect identification of the branch, and lack of the proper velocity structure at the velocity point, are avoided and the analysis shows that one-way travel of P waves through oceanic mantle is delayed by 0.65 to 0.95 sec relative to United States mid-continental mantle.

A new P-wave velocity core model is constructed from observed times, dt/dΔ's, and relative amplitudes of P'; the observed times of SKS, SKKS, and PKiKP; and a new mantle-velocity determination by Jordan and Anderson. The new core model is smooth except for a discontinuity at the inner-core boundary determined to be at a radius of 1215 km. Short-period amplitude data do not require the inner core Q to be significantly lower than that of the outer core. Several lines of evidence show that most, if not all, of the arrivals preceding the DF branch of P' at distances shorter than 143° are due to scattering as proposed by Haddon and not due to spherically symmetric discontinuities just above the inner core as previously believed. Calculation of the travel-time distribution of scattered phases and comparison with published data show that the strongest scattering takes place at or near the core-mantle boundary close to the seismic station.

In Part II, the largest events in the San Fernando earthquake series, initiated by the main shock at 14 00 41.8 GMT on February 9, 1971, were chosen for analysis from the first three months of activity, 87 events in all. The initial rupture location coincides with the lower, northernmost edge of the main north-dipping thrust fault and the aftershock distribution. The best focal mechanism fit to the main shock P-wave first motions constrains the fault plane parameters to: strike, N 67° (± 6°) W; dip, 52° (± 3°) NE; rake, 72° (67°-95°) left lateral. Focal mechanisms of the aftershocks clearly outline a downstep of the western edge of the main thrust fault surface along a northeast-trending flexure. Faulting on this downstep is left-lateral strike-slip and dominates the strain release of the aftershock series, which indicates that the downstep limited the main event rupture on the west. The main thrust fault surface dips at about 35° to the northeast at shallow depths and probably steepens to 50° below a depth of 8 km. This steep dip at depth is a characteristic of other thrust faults in the Transverse Ranges and indicates the presence at depth of laterally-varying vertical forces that are probably due to buckling or overriding that causes some upward redirection of a dominant north-south horizontal compression. Two sets of events exhibit normal dip-slip motion with shallow hypocenters and correlate with areas of ground subsidence deduced from gravity data. Several lines of evidence indicate that a horizontal compressional stress in a north or north-northwest direction was added to the stresses in the aftershock area 12 days after the main shock. After this change, events were contained in bursts along the downstep and sequencing within the bursts provides evidence for an earthquake-triggering phenomenon that propagates with speeds of 5 to 15 km/day. Seismicity before the San Fernando series and the mapped structure of the area suggest that the downstep of the main fault surface is not a localized discontinuity but is part of a zone of weakness extending from Point Dume, near Malibu, to Palmdale on the San Andreas fault. This zone is interpreted as a decoupling boundary between crustal blocks that permits them to deform separately in the prevalent crustal-shortening mode of the Transverse Ranges region.

Algebraic Techniques in Coding Theory: Entropy Vectors, Frames, and Constrained Coding

The study of codes, classically motivated by the need to communicate information reliably in the presence of error, has found new life in fields as diverse as network communication, distributed storage of data, and even has connections to the design of linear measurements used in compressive sensing. But in all contexts, a code typically involves exploiting the algebraic or geometric structure underlying an application. In this thesis, we examine several problems in coding theory, and try to gain some insight into the algebraic structure behind them.

The first is the study of the entropy region - the space of all possible vectors of joint entropies which can arise from a set of discrete random variables. Understanding this region is essentially the key to optimizing network codes for a given network. To this end, we employ a group-theoretic method of constructing random variables producing so-called "group-characterizable" entropy vectors, which are capable of approximating any point in the entropy region. We show how small groups can be used to produce entropy vectors which violate the Ingleton inequality, a fundamental bound on entropy vectors arising from the random variables involved in linear network codes. We discuss the suitability of these groups to design codes for networks which could potentially outperform linear coding.

The second topic we discuss is the design of frames with low coherence, closely related to finding spherical codes in which the codewords are unit vectors spaced out around the unit sphere so as to minimize the magnitudes of their mutual inner products. We show how to build frames by selecting a cleverly chosen set of representations of a finite group to produce a "group code" as described by Slepian decades ago. We go on to reinterpret our method as selecting a subset of rows of a group Fourier matrix, allowing us to study and bound our frames' coherences using character theory. We discuss the usefulness of our frames in sparse signal recovery using linear measurements.

The final problem we investigate is that of coding with constraints, most recently motivated by the demand for ways to encode large amounts of data using error-correcting codes so that any small loss can be recovered from a small set of surviving data. Most often, this involves using a systematic linear error-correcting code in which each parity symbol is constrained to be a function of some subset of the message symbols. We derive bounds on the minimum distance of such a code based on its constraints, and characterize when these bounds can be achieved using subcodes of Reed-Solomon codes.

Assessment of existing information on Atlantic coastal fish habitats: development of a web-based spatial bibliography, query tools, and data summaries

The primary objective of this project, “the Assessment of Existing Information on Atlantic Coastal Fish Habitat”, is to inform conservation planning for the Atlantic Coastal Fish Habitat Partnership (ACFHP). ACFHP is recognized as a Partnership by the National Fish Habitat Action Plan (NFHAP), whose overall mission is to protect, restore, and enhance the nation’s fish and aquatic communities through partnerships that foster fish habitat conservation. This project is a cooperative effort of NOAA/NOS Center for Coastal Monitoring and Assessment (CCMA) Biogeography Branch and ACFHP. The Assessment includes three components; 1. a representative bibliographic and assessment database, 2. a Geographical Information System (GIS) spatial framework, and 3. a summary document with description of methods, analyses of habitat assessment information, and recommendations for further work. The spatial bibliography was created by linking the bibliographic table developed in Microsoft Excel and exported to SQL Server, with the spatial framework developed in ArcGIS and exported to GoogleMaps. The bibliography is a comprehensive, searchable database of over 500 selected documents and data sources on Atlantic coastal fish species and habitats. Key information captured for each entry includes basic bibliographic data, spatial footprint (e.g. waterbody or watershed), species and habitats covered, and electronic availability. Information on habitat condition indicators, threats, and conservation recommendations are extracted from each entry and recorded in a separate linked table. The spatial framework is a functional digital map based on polygon layers of watersheds, estuarine and marine waterbodies derived from NOAA’s Coastal Assessment Framework, MMS/NOAA’s Multipurpose Marine Cadastre, and other sources, providing spatial reference for all of the documents cited in the bibliography. Together, the bibliography and assessment tables and their spatial framework provide a powerful tool to query and assess available information through a publicly available web interface. They were designed to support the development of priorities for ACFHP’s conservation efforts within a geographic area extending from Maine to Florida, and from coastal watersheds seaward to the edge of the continental shelf. The Atlantic Coastal Fish Habitat Partnership has made initial use of the Assessment of Existing Information. Though it has not yet applied the AEI in a systematic or structured manner, it expects to find further uses as the draft conservation strategic plan is refined, and as regional action plans are developed. It also provides a means to move beyond an “assessment of existing information” towards an “assessment of fish habitat”, and is being applied towards the National Fish Habitat Action Plan (NFHAP) 2010 Assessment. Beyond the scope of the current project, there may be application to broader initiatives such as Integrated Ecosystem Assessments (IEAs), Ecosystem Based Management (EBM), and Marine Spatial Planning (MSP).

Identification of Prorocentrum minimum and Takayama pulchella by fluorescence in situ hybridization through epifluorescence microscopy and flow cytometry

Partial rDNA sequences of Prorocentrum minimum and Takayama pulchella were amplified, cloned and sequenced. and these sequence data were deposited in the GenBank. Eight oligonucleotide probes (DNA probes) were designed based on the sequence analysis. The probes were employed to detect and identify P. minimum and T. pulchella in unialgal and mixed algal samples with a fluorescence in situ hybridization method using flow cytometry. Epifluorescence micrographs showed that these specific probes labeled with fluorescein isothiocyanate entered the algal cells and bound to target sequences, and the fluorescence signal resulting from whole-cell hybridization varied from probe to probe. These DNA probes and the hybridization protocol we developed were specific and effective for P. minimum and T. pulchella, without any specific binding to other algal species. The hybridization efficiency of different probes specific to P. minimum was in the order: PM18S02 > PM28S02 > PM28S01 > PM18S01, and that of the probes specific to T. pulchella was TP18S02 > TP28S01 > TP28S02 > TP18S01. The different hybridization efficiency of the DNA probes could also be shown in the fluorescent signals between the labeled and unlabeled cells demonstrated using flow cytometry. The DNA probes PM18S02, PM28S02; TP18S02 and TP28S01, and the protocol, were also useful for the detection of algae in natural samples.

Intrasubband and intersubband transitions in lightly and heavily doped GaAs/AlGa1-xAs multiple quantum wells

We use a polarizer to investigate quantum-well infrared absorption, and report experimental results as follows. The intrasubband transition was observed in GaAs/AlxGa1-xAs multiple quantum wells (MQWs) when the incident infrared radiation (IR) is polarized parallel to the MQW plane. According to the selection rule, an intrasubband transition is forbidden. Up to now, most studies have only observed the intersubband transition between two states with opposite parity. However, our experiment shows not only the intersubband transitions, but also the intrasubband transitions. In our study, we also found that for light doping in the well (4x10(18) cm(-3)), the intrasubband transition occurs only in the lowest subband, while for the heavy doping (8x10(18) cm(-3)), such a transition occurs not only in the lowest subband, but also in the first excited one, because of the electron subband filling. Further experimental results show a linear dependence of the intrasubband transition frequency on the root of the well doping density. These data are in good agreement with our numerical results. Thus we strongly suggest that such a transition can be attributed to plasma oscillation. Conversely, when the incident IR is polarized perpendicular to the MQW plane, intersubband-transition-induced signals appear, while the intrasubband-transition-induced spectra disappear for both light and heavy well dopings. A depolarization blueshift was also taken into account to evaluate the intersubband transition spectra at different well dopings. Furthermore, we performed a deep-level transient spectroscopy (DLTS) measurement to determine the subband energies at different well dopings. A good agreement between DLTS, infrared absorption, and numerical calculation was obtained. In our experiment, two important phenomena are noteworthy: (1) The polarized absorbance is one order of magnitude higher than the unpolarized spectra. This puzzling result is well explained in detail. (2) When the IR, polarized perpendicular to the well plane, normally irradiates the 45 degrees-beveled edge of the samples, we only observed intersubband transition spectra. However, the intrasubband transition signals caused by the in-plane electric-field component are significantly absent. The reason is that such in-plane electric-field components can cancel each other out everywhere during the light propagating in the samples. The spectral widths of bound-to-bound and bound-to-continuum transitions were also discussed, and quantitatively compared to the relaxation time tau, which is deduced from the electron mobility. The relaxation times deduced from spectral widths of bound-to-bound and bound-to-continuum transitions are also discussed, and quantitatively compared to the relaxation time deduced from electron mobility. [S0163-1829(98)01912-2].

Superluminescent diode monolithically integrated with novel Y-branch by bundle integrated waveguide for fiber optic gyroscope - art. no. 68380D

We have developed a novel InP-based, ridge-waveguide photonic integrated circuit (PIC), which consists of a 1.1-um wavelength Y-branch optical waveguide with low loss and improved far field pattern and a 1.3-um wavelength strained InGaAsP-InP multiple quantum-well superluminescent diode, with bundle integrated guide (BIG) as the scheme for monolithic integration. The simulations of BIG and Y-branches show low losses and improved far-field patterns, based on the beam propagation method (BPM). The amplified spontaneous emission of the device is up to 10 mW at 120 mA with no threshold and saturation. Spectral characteristics of about 30 nm width and less than I dB modulation are achieved using the built-in anti-lasing ability of Y-branch. The beam divergence angles in horizontal and vertical directions are optimized to as small as 12 degrees x8 degrees, resulting in good fiber coupling. The compactness, simplicity in fabrication, good superluminescent performance, low transmission loss and estimated low coupling loss prove the BIG and Y-branch method to be a feasible way for integration and make the photonic integrated circuit of Y-branch and superluminescent diode an promising candidate for transmitter and transceiver used in fiber optic gyroscope.

A voltage-controlled tunable two-color infrared photodetector using GaAs/AlAs/GaAlAs and GaAs/GaAlAs stacked multiquantum wells

A voltage-controlled tunable two-color infrared detector with photovoltaic (PV) and photoconductive (PC) dual-mode operation at 3-5 mu m and 8-14 mu m using GaAs/AlAs/AlGaAs double barrier quantum wells (DBQWs) and bound-to-continuum GaAs/AlGaAs quantum wells is demonstrated. The photoresponse peak of the photovoltaic GaAs/AlAs/GaAlAs DBQWs is at 5.3 mu m, and that of the photoconductive GaAs/GaAlAs quantum wells is at 9.0 mu m. When the two-color detector is under a zero bias, the spectral response at 5.3 mu m is close to saturate and the peak detectivity at 80 K can reach 1.0X10(11) cmHz(1/2)/W, while the spectral photoresponsivity at 9.0 mu m is absolutely zero completely. When the external voltage of the two-color detector is changed to 2.0 V, the spectral photoresponsivity at 5.3 mu m becomes zero while the spectral photoresponsivity at 9.0 mu m increases comparable to that at 5.3 mu m under zero bias, and the peak detectivity (9.0 mu m) at 80 K can reach 1.5X10(10) cmHz(1/2)/W. Strictly speaking, this is a real bias-controlled tunable two-color infrared photodetector. We have proposed a model based on the PV and PC dual-mode operation of stacked two-color QWIPs and the effects of tunneling resonance with narrow energy width of photoexcited electrons in DBQWs, which can explain qualitatively the voltage-controlled tunable behavior of the photoresponse of the two-color infrared photodetector. (C) 1996 American Institute of Physics.

Model atmosphere and kinematical analyses of early-type stars from the Edinburgh-Cape Survey

We present high-resolution spectroscopic observations of 21 B- type stars, selected from the Edinburgh-Cape Blue Object Survey. Model atmosphere analyses confirm that 14 of these stars are young, main-sequence B-type objects with Population I chemical compositions. The remaining seven are found to be evolved objects, including subdwarfs, horizontal branch and post-AGB objects. A kinematical analysis shows that all 14 young main-sequence stars could have formed in the disc and subsequently been ejected into the halo. These results are combined with the analysis of a previous subsample of stars taken from the Survey. Of the complete sample, 31 have been found to be young, main-sequence objects, with formation in the disc, and subsequent ejection into the halo, again being found to be a plausible scenario.

High-resolution spectroscopy of globular cluster post-Asymptotic Giant Branch stars

We present model atmosphere analyses of high resolution Keck and VLT optical spectra for three evolved stars in globular clusters, viz. ZNG-1 in M 10, ZNG-1 in M 15 and ZNG-1 in NGC 6712. The derived atmospheric parameters and chemical compositions confirm the programme stars to be in the post- Asymptotic Giant Branch (post-AGB) evolutionary phase. Differential abundance analyses reveal CNO abundance patterns in M 10 ZNG-1, and possibly M 15 ZNG-1, which Suggest that both objects may have evolved off the AGB before the third dredge-up occurred. The abundance pattern of these stars is similar to the third class of optically, bright post-AGB objects discussed by van Winckel (1997). Furthermore, M 10 ZNG-1 exhibits a large C underabundance (with Delta[C/O] similar to -1.6 dex), typical of other hot post-AGB objects. Differential Delta[alpha/Fe] abundance ratios in both M 10 ZNG-1 and NGC 6712 ZNG-1 are found to be approximately 0.0 dex, with the Fe abundance of the former being in disagreement with the cluster metallicity of M 10. Given that the Fe absorption features in both M 10 ZNG-1 and NGC6712 ZNG-1 are well observed and reliably modelled, we believe that these differential Fe abundance estimates to be secure. However, our Fe abundance is difficult to explain in terms of previous evolutionary processes that Occur oil both the Horizontal Branch and the AGB.

THE ULTRASTRUCTURE AND IMMUNOGOLD LABELING OF PANCREATIC POLYPEPTIDE-IMMUNOREACTIVE CELLS ASSOCIATED WITH THE EGG-FORMING APPARATUS OF A MONOGENEAN PARASITE, DICLIDOPHORA-MERLANGI

An electron microscopical examination has been made of the fine structure and disposition of pancreatic polypeptide immunoreactive cells associated with the egg-forming apparatus in Diclidophora merlangi. The cell bodies are positioned in the parenchyma surrounding the ootype and taper to axon-like processes that extend to the ootype wall. The terminal regions of these processes branch and anastomose and, in places, the swollen endings or varicosities form synaptic appositions with the muscle fibres in the ootype wall. The cells are characterized by an extensive GER-Golgi system that is involved in the assembly and packaging of dense-cored vesicles. The vesicles accumulate in the axons and terminal varicosities, and their contents were found to be immunoreactive with antisera raised to the C-terminal hexapeptide amide of pancreatic polypeptide. It is concluded that the cells are neurosecretory in appearance and that, functionally, their secretions may serve to regulate ootype motility and thereby help co-ordinate egg production in the worm.

Partial and total multiphoton detachment rates for H- in a perturbative B-spline-based approach

An ab initio approach has been applied to study multiphoton detachment rates for the negative hydrogen ion in the lowest nonvanishing order of perturbation theory. The approach is based on the use of B splines allowing an accurate treatment of the electronic repulsion. Total detachment rates have been determined for two- to six-photon processes as well as partial rates for detachment into the different final symmetries. It is shown that B-spline expansions can yield accurate continuum and bound-state wave functions in a very simple manner. The calculated total rates for two- and three-photon detachment are in good agreement with other perturbative calculations. For more than three-photon detachment little information has been available before now. While the total cross sections show little structure, a fair amount of structure is predicted in the partial cross sections. In the two-photon process, it is shown that the detached electrons mainly have s character. For four- and six-photon processes, the contribution from the d channel is the most important. For three- and five-photon processes p electrons dominate the electron emission spectrum. Detachment rates for s and p electrons show minima as a function of photon energy. © 1994 The American Physical Society.

Degradation of 4-fluorobiphenyl in soil investigated by 19F NMR spectroscopy and 14C radiolabelling analysis

The incubation of the model pollutant [U-14C]'-4-fluorobiphenyl (4FBP) in soil, in the presence and absence of biphenyl (a co-substrate), was carried out in order to study the qualitative disposition and fate of the compound using 14C-HPLC and 19F NMR spectroscopy. Components accounted for using the radiolabel were volatilization, CO2 evolution, organic solvent extractable and bound residue. Quantitative analysis of these data gave a complete mass balance. After sample preparation. 14C-HPLC was used to establish the number of 4FBP related components present in the organic solvent extract. 19F NMR was also used to quantify the organic extracts and to identify the components of the extract. Both approaches showed that the composition of the solvent extractable fractions comprised only parent compound with no metabolites present. As the 14C radiolabel was found to be incorporated into the soil organic matter this indicates that metabolites were being generated, but were highly transitory as incorporation into the SOM was rapid. The inclusion of the co-substrate biphenyl was to increase the overall rate of degradation of 4FBP in soil. The kinetics of disappearance of parent from the soil using the data obtained were investigated from both techniques. This is the first report describing the degradation of a fluorinated biphenyl in soil.