DNA sequencing and genotyping by transcriptional synthesis of chain-terminated RNA ladders and MALDI-TOF mass spectrometry

Sets of RNA ladders can be synthesized by transcription of a bacteriophage-encoded RNA polymerase using 3′-deoxynucleotides as chain terminators. These ladders can be used for sequencing of DNA. Using a nicked form of phage SP6 RNA polymerase in this study substantially enhanced yields of transcriptional sequencing ladders. Matrix-assisted laser desorption/ionization time-of-flight mass spectrometry (MALDI-TOF MS) of chain-terminated RNA ladders allowed DNA sequence determination of up to 56 nt. It is also demonstrated that A→G and C→T variations in heterozygous and homozygous samples can be unambiguously identified by the mass spectrometric analysis. As a step towards single-tube sequencing reactions, α-thiotriphosphate nucleotide analogs were used to overcome problems caused by chain terminator-independent, premature termination and by the small mass difference between natural pyrimidine nucleotides.

Rapid mass spectrometric peptide sequencing and mass matching for characterization of human melanoma proteins isolated by two-dimensional PAGE.

We report a general mass spectrometric approach for the rapid identification and characterization of proteins isolated by preparative two-dimensional polyacrylamide gel electrophoresis. This method possesses the inherent power to detect and structurally characterize covalent modifications. Absolute sensitivities of matrix-assisted laser desorption ionization and high-energy collision-induced dissociation tandem mass spectrometry are exploited to determine the mass and sequence of subpicomole sample quantities of tryptic peptides. These data permit mass matching and sequence homology searching of computerized peptide mass and protein sequence data bases for known proteins and design of oligonucleotide probes for cloning unknown proteins. We have identified 11 proteins in lysates of human A375 melanoma cells, including: alpha-enolase, cytokeratin, stathmin, protein disulfide isomerase, tropomyosin, Cu/Zn superoxide dismutase, nucleoside diphosphate kinase A, galaptin, and triosephosphate isomerase. We have characterized several posttranslational modifications and chemical modifications that may result from electrophoresis or subsequent sample processing steps. Detection of comigrating and covalently modified proteins illustrates the necessity of peptide sequencing and the advantages of tandem mass spectrometry to reliably and unambiguously establish the identity of each protein. This technology paves the way for studies of cell-type dependent gene expression and studies of large suites of cellular proteins with unprecedented speed and rigor to provide information complementary to the ongoing Human Genome Project.

The evolution and masses of the neutron star and donor star in the high mass X-ray binary OAO 1657−415

We report near-infrared radial velocity (RV) measurements of the recently identified donor star in the high mass X-ray binary (HMXB) system OAO 1657−415 obtained in the H band using ISAAC on the Very Large Telescope. Cross-correlation methods were employed to construct a RV curve with a semi-amplitude of 22.1 ± 3.5 km s−1. Combined with other measured parameters of this system it provides a dynamically determined neutron star (NS) mass of 1.42 ± 0.26 M⊙ and a mass of 14.3 ± 0.8 M⊙ for the Ofpe/WN9 highly evolved donor star. OAO 1657−415 is an eclipsing HMXB pulsar with the largest eccentricity and orbital period of any within its class. Of the 10 known eclipsing X-ray binary pulsars OAO 1657−415 becomes the ninth with a dynamically determined NS mass solution and only the second in an eccentric system. Furthermore, the donor star in OAO 1657−415 is much more highly evolved than the majority of the supergiant donors in other HMXBs, joining a small but growing list of HMXBs donors with extensive hydrogen depleted atmospheres. Considering the evolutionary development of OAO 1657−415, we have estimated the binding energy of the envelope of the mass donor and find that there is insufficient energy for the removal of the donor’s envelope via spiral-in, ruling out a common envelope evolutionary scenario. With its non-zero eccentricity and relatively large orbital period the identification of a definitive evolutionary pathway for OAO 1657−415 remains problematic, we conclude by proposing two scenarios which may account for OAO 1657−415 current orbital configuration.

An XMM-Newton view of FeKα in high-mass X-ray binaries

We present a comprehensive analysis of the whole sample of available XMM-Newton observations of high-mass X-ray binaries (HMXBs) until August 2013, focusing on the FeKα emission line. This line is key to better understanding the physical properties of the material surrounding the X-ray source within a few stellar radii (the circumstellar medium). We collected observations from 46 HMXBs and detected FeKα in 21 of them. We used the standard classification of HMXBs to divide the sample into different groups. We find that (1) different classes of HMXBs display different qualitative behaviours in the FeKα spectral region. This is visible especially in SGXBs (showing ubiquitous Fe fluorescence but not recombination Fe lines) and in γ Cass analogues (showing both fluorescent and recombination Fe lines). (2) FeKα is centred at a mean value of 6.42 keV. Considering the instrumental and fits uncertainties, this value is compatible with ionization states that are lower than Fe xviii. (3) The flux of the continuum is well correlated with the flux of the line, as expected. Eclipse observations show that the Fe fluorescence emission comes from an extended region surrounding the X-ray source. (4) We observe an inverse correlation between the X-ray luminosity and the equivalent width of FeKα (EW). This phenomenon is known as the X-ray Baldwin effect. (5) FeKα is narrow (σline< 0.15 keV), reflecting that the reprocessing material does not move at high speeds. We attempt to explain the broadness of the line in terms of three possible broadening phenomena: line blending, Compton scattering, and Doppler shifts (with velocities of the reprocessing material V ~ 1000 km s-1). (6) The equivalent hydrogen column (NH) directly correlates to the EW of FeKα, displaying clear similarities to numerical simulations. It highlights the strong link between the absorbing and the fluorescent matter. (7) The observed NH in supergiant X-ray binaries (SGXBs) is in general higher than in supergiant fast X-ray transients (SFXTs). We suggest two possible explanations: different orbital configurations or a different interaction compact object – wind. (8) Finally, we analysed the sources IGR J16320-4751 and 4U 1700-37 in more detail, covering several orbital phases. The observed variation in NH between phases is compatible with the absorption produced by the wind of their optical companions. The results clearly point to a very important contribution of the donor’s wind in the FeKα emission and the absorption when the donor is a supergiant massive star.

Characterization of rock slopes through slope mass rating using 3D point clouds

Rock mass classification systems are widely used tools for assessing the stability of rock slopes. Their calculation requires the prior quantification of several parameters during conventional fieldwork campaigns, such as the orientation of the discontinuity sets, the main properties of the existing discontinuities and the geo-mechanical characterization of the intact rock mass, which can be time-consuming and an often risky task. Conversely, the use of relatively new remote sensing data for modelling the rock mass surface by means of 3D point clouds is changing the current investigation strategies in different rock slope engineering applications. In this paper, the main practical issues affecting the application of Slope Mass Rating (SMR) for the characterization of rock slopes from 3D point clouds are reviewed, using three case studies from an end-user point of view. To this end, the SMR adjustment factors, which were calculated from different sources of information and processes, using the different softwares, are compared with those calculated using conventional fieldwork data. In the presented analysis, special attention is paid to the differences between the SMR indexes derived from the 3D point cloud and conventional field work approaches, the main factors that determine the quality of the data and some recognized practical issues. Finally, the reliability of Slope Mass Rating for the characterization of rocky slopes is highlighted.

Multiwavelength study of the fast rotating supergiant high-mass X-ray binary IGR J16465−4507

Context. Since its launch, the X-ray and γ-ray observatory INTEGRAL satellite has revealed a new class of high-mass X-ray binaries (HMXB) displaying fast flares and hosting supergiant companion stars. Optical and infrared (OIR) observations in a multi-wavelength context are essential to understand the nature and evolution of these newly discovered celestial objects. Aims. The goal of this multiwavelength study (from ultraviolet to infrared) is to characterise the properties of IGR J16465−4507, to confirm its HMXB nature and that it hosts a supergiant star. Methods. We analysed all OIR, photometric and spectroscopic observations taken on this source, carried out at ESO facilities. Results. Using spectroscopic data, we constrained the spectral type of the companion star between B0.5 and B1 Ib, settling the debate on the true nature of this source. We measured a high rotation velocity of v = 320 ± 8km s-1 from fitting absorption and emission lines in a stellar spectral model. We then built a spectral energy distribution from photometric observations to evaluate the origin of the different components radiating at each energy range. Conclusions. We finally show that, having accurately determined the spectral type of the early-B supergiant in IGR J16465−4507, we firmly support its classification as an intermediate supergiant fast X-ray transient (SFXT).

An account of my quarter bills, charges &c from July 11th 1743 to July 13th 1748 at Harvard College

Handwritten account book kept while Storer was a student at Harvard College. The well-organized volume is arranged by expense type and then date and was updated periodically, usually quarterly. The information offers a glimpse at the expenses of a Harvard student and provides information about the larger community that supported student life. The precise entries indicate the lifelong habits of Storer as a careful and methodical financial manager that would prove so valuable when he served as Harvard's treasurer more than thirty years later. Storer documents accounts with the steward, butler, sweeper, glazier, barber, and lists these individuals by name. The volume also includes notes on expenses for boarding, transportation, wood, and pocket expenses. While most entries do not list specific purchases, Storer provides details on the cost of a Harvard Commencement in 1747 (including the cost of a diploma, money to the President, hiring a house, a boat, a woman, and "2 Negroes"), and a specific accounting of the different food purchased for the event; Storer also lists expenses for an 1748 "supper for the graduates."

Manuscript copy of the book of Harvard, 1767

Handwritten copy of the Book of Harvard written on one large sheet of paper and signed Boston, January 10th, 1767.

Annual report of the Secretary of the Board of Agriculture.

26

Annual report of the Secretary of the Board of Agriculture.

28

Annual report of the Secretary of the Board of Agriculture.

4

Annual report of the Secretary of the Board of Agriculture.

7

Annual report of the Secretary of the Board of Agriculture.

20

Annual report of the Secretary of the Board of Agriculture.

23

Annual report of the Secretary of the Board of Agriculture.

3