dataesr/scientific-paragraphs-categorization huggingface dataset:
Estimation of microsatellite loci success rates for the different species To estimate the number of PALs based on the numbers of polymorphic loci initially identified, we used steps A through I described below. (A) The PHOBOS software version 3. 3. 11 (Mayer 2007 ) was used to assign obtained sequence reads to a target species on the basis of the species-specific MID tags. PHOBOS was also used to identify sequence reads containing noninterrupted stretches of at least ten tetra-nucleotide repeat motifs. (B) Selected sequences were retained by PHOBOS only when the flanking region on each side of the repeat motif was at least 25 bp long. These sequences were then assessed by eye for their general suitability for primer design, that is, sequences with more than five repetitive nucleotides at a stretch were removed.
2002). Loci are grouped by the multiplex combinations used for amplification. Information on the locus name, primer sequence, direction (F is forward, R is reverse), annealing temperature of the primer for PCRs, microsatellite motif, amplified fragment size range, number of alleles, and labeling dye are provided together with the accession number of the associated GenBank sequence. based microsatellite loci development in the meadow viper resulted in only 14 applicable loci out of 37,000 sequence reads (0. 037% success rate) and in a success rate of only 0. 007% in the Asp viper  (Geser et al. 2013 )both studies were performed without prior enrichment of genomic sublibraries. Accordingly, our obtained success rates are comparable quite high and seem to justify the applied enrichment procedure.
Locus name There is no doubt that Illumina sequencing is by far more cost-effective than 454-sequencing. Castoe et al. (2012b)  suggest that Illumina-based microsatellite loci development is by far more effective than 454-based Table  4. Characterization of the full set of 20 applied microsatellite loci for Calotriton asper. Loci are grouped by multiplex combinations used for amplification. Locus name, primer sequence, direction (F is forward, R is reverse), annealing temperature of the primer for PCRs, microsatellite motif, amplified fragment size range, number of alleles, labeling dye, and GenBank accession number are provided. The number of alleles of C. asper microsatellite loci detected in C. arnoldi cross-amplification is also provided; polymorphic loci are highlighted in bold. approaches.
Acknowledgments We thank  A. Funke  ( Untere Landschaftsbeho ¨rde Krefeld Germany ), the  Pr efecture de la R egion Languedoc Roussillon (Gard and H erault, France) , the  DREAL de l'Ari ege and Haute Garonne (France) ,  Gobierno de Aragon (Spain)  and  Generalitat de Catalunya (Spain)  for providing collection permits. R. Barral, J. M. and  M. G. Buresi ,  D. Desfe  `re, and  H. Mialane  kindly provided access to their private ponds. F. Amat ,  S. Carranza ,  D. Ortmann ,  D. Sanuy ,  L. Winandy  and  J er emie Cornuau  assisted during fieldwork. S. Hauswaldt  ( Technical University of Braunschweig ) and  T. C. Glenn  ( University of Georgia ) provided helpful suggestions on the microsatellite loci enrichment and sequencing. This research has been funded by the  Deutsche Bundesstiftung Umwelt (DBU)  through a Ph.
1%, Triton X-100, 0. 5 mg/ml Pepstatine and 1X to 3X HaltTM protease/ hosphatase inhibitors; Thermo Scientific) was used. Cells were left on ice during 5 min then spun 3 min at 7500 r. p. m. at 41C. Pellets were incubated in solution A for 5 min on ice, spun again, and resuspended in solution A (Chromatin fraction). The supernatants from the two extraction steps were pooled (soluble fraction). Chromatin fractions were sonicated. Whole cell extracts were performed as previously described  (Betous et al, 2009). Blots were detected by ECL Western Blotting Substrate (Pierce) or ECL Plus Western blot detecting system (GE Healthcare). Where indicated, western blots were quantitated using ImageJ software.
Microscope image acquisition Images acquisition of multiple random fields were carried out on a wide field Leica DMLA microscope equipped with Â 63 oil immersion objective (PLAPO 1. 4 Leica) using a Cool-Snap HQ CCD camera (Photometrics, Roper Scientific) driven by PM capture Pro6. 0 or Metamorph software. Images were assembled with Adobe photoshop and Illustrator. When indicated in the figure legends, images of RPA were acquired using the Â 100 NA 1. 4 objective of a SP2 confocal microscope (Leica microsytems) using laser lines 488 and 633 nm for excitation of Alexa Fluor 488 and ToPro3 dyes.
Cell-cycle analysis and measurement by flow cytometry In all, 100 000 cells were seeded 48 h prior to harvesting. Cells were washed in PBS once and 300 ml of Propidium Iodide solution (25 mg/ml Propidium Iodide (Sigma), 0. 1% C 6 H 5 O 7 Na 3 , 2H 2 O, 10% RNAse A (Sigma), 0. 1% Triton X-100) was added 30 min prior to FACS analysis (FACScan, Becton Dickinson). Results were analysed by using Modfit software. Analysis g-H2AX and H3P by flow cytometry were performed as described  (Betous et al, 2009), H3P (Cell Signaling, 1:75).
Supplementary data Supplementary data are available at The EMBO Journal Online (http://www. embojournal. org). and YPOL-2010 to JSH), 1 ARC grant (3156 to DM), and FRM (FRM Equipes to DM). Author contributions: RB, MJP, DM and JSH designed the experiments. RB, MJP, LP, SVdL and EOS performed the experiments in human cells. BR, MJP and DM performed the experiments in Xenopus extracts. NN, PG and TN performed the production of the human Pol Kappa antibodies. RB, MJP, CG, EF, CC, DM and JSH analysed the data and contributed to the manuscript preparation. DM and JSH supervised the project and wrote the manuscript.
Acknowledgements We thank  Renaud Poincloux  and  Franc¸oise Viala  ( IPBS, CNRS Toulouse ) for their technical contributions. We thank  S Waga  ( Japan Women's University ) for anti Xpol d antibodies. This work was supported by  La Ligue  contre le cancer (Equipe Labellisee 2013 to JSH; PhD grant for RB), 2  INCa  Grants (CHECKPOL-2007.
Cell viability is not compromised as there is no chromatin damage. Dbait32Hc treatment is sensitizing tumors to radiotherapy and is currently under clinical investigation for local metastatic melanoma treatment (clinicaltrials. gov/show/NCT01469455). Therefore, it is particularly important to identify phosphorylated targets of DNA-PK after Dbait32Hc treatment as eventual activity biomarkers. With the aim of identifying new downstream targets of DNA-PK, we used Dbait32Hc to investigate the impact of DNA-PK activation on the phosphoproteome, in a two-dimensional gel electrophoresis (2DE)-based approach. We observed an increase in the phosphorylation of proteins involved in cell signaling, protein stability and degradation and several proteins involved in the cytoskeleton. To our best knowledge a relationship between DNA-PK signaling and the cytoskeleton has never been described.
The mass spectrometer was set to acquire a single MS scan followed by up to five data-dependent scans and, if a neutral loss of 98 Da from the precursor ion was observed in the CID mass spectrum, an MS3 scan of the neutral loss ion was also carried out (simultaneous fragmentation of neutral loss product and precursor, dynamic exclusion repeat count of 1, repeat duration of 30 seconds, exclusion duration of 180 seconds and lock-mass option enabled). The resulting spectra were then analyzed with Mascot TM and SEQUESTH Software (Matrix Science and Thermo Fisher Scientific), created with Proteome Discoverer (version: 1. 2. 0. 92, Thermo Fisher Scientific), using the NCBInr Homo sapiens (human 2011 05 03, 236201 sequences) protein database. All phosphorylated peptides with unphosphorylated counterparts were validated by hand.
For immunofluorescence staining, cells were processed as previously described  [18]. Microscopy was performed at room temperature with the Leica SP5 confocal system, attached to a DMI6000 stand, with a 636/1. 4 oil immersion objective. Images were processed with the freely available ImageJ software (http:// rsb. info. nih. gov. gate1. inist. fr/ij/) and the LOCI bioformat plug-in (http://www. loci. wisc. edu/ome/formats. html), to access images generated by the Leica SP5 confocal system. For immunoblotting, cells were lysed by scraping into Laemmli buffer and boiling for 10 min. The resulting lysates were then centrifuged and protein levels were normalized with the BCA protein assay kit (Thermo Scientific, USA). Proteins were separated by SDS-PAGE in 12% polyacrylamide  (35. 5  acrylamide/1 bisacrylamide) gels, transferred to nitrocellulose membranes, blocked by incubation with Odyssey buffer (LI-COR Biosciences, Lincoln, NE, USA) for 1 h and hybridized overnight at 4uC with primary antibody diluted in Odyssey buffer.
For Western blots, the membranes were probed with goat anti-mouse or anti-rabbit secondary antibodies conjugated to Alexa Fluor 680 (Invitrogen) or IRdye 800 (Rockland Immunochemicals, Gilbertsville, PA, USA). The blots were imaged and quantified with the Odyssey Infrared Imaging System (LI-COR Biosciences, Lincoln) and Odyssey software. For detection of MOS protein the nitrocellulose membranes were blocked with 5% nonfat milk overnight at 4uC and hybridized also overnight at 4uC with primary antibody. Blots were then incubated with horseradish peroxidase conjugated goat anti-rabbit IgG secondary antibodies (sc-2004, Santa Cruz Biotechnology, Inc, Germany) and protein-antibody complexes were revealed on amersham hyperfilm (GE Healthcare, UK). Cell cycle analysis was performed by flow cytometry, according to standard protocols.
Briefly, the cells were fixed by incubation in ice-cold ethanol for 18 h, permeabilized by incubation with 0. 2 % Triton X-100 for 15 min, treated with 25 U/ml RNAse A (Invitrogen) and DNA was stained with 50 mg/ml propidium iodide (Sigma-Aldrich). Cells were analyzed with a FACScalibur flow cytometer (BD Biosciences, Franklin Lakes, NJ, USA) and data were analyzed with the freely available WinMDI 2. 8 (Scripps Research Institute, La Jolla, CA, USA) software.
Wound healing assay and adhesion assay Cells were cultured as monolayers in six-well plates until confluence and were then wounded with a yellow pipette tip. At least three wounds (in duplicate) were created for each set of experimental conditions, and the experiment was repeated two or three times. Wound closure was monitored in phase contrast mode, with a 4/5D microscope. This system consisted of a Leica DM IRBE microscope (Leica Microsystems, France) equipped with an incubation chamber (37uC, humidified atmosphere containing 5% CO 2 ), an x-y-z stage controller and a chargecoupled device (CCD) CoolSNAP HQ2 camera (Photometrics, Tucson, AZ). Images were acquired at 15-minute intervals over a period of 16 h, with Metamorph software (Molecular Devices, Sunnyvale, CA). Movies were then reconstructed with the ''nd stack builder'', plugins from ImageJ software (http://rsbweb.
nih. gov/ij/plugins/track/builder. html) developed by F. Cordelieres at Institut Curie (Orsay, France). Wound closure area was calculated manually with ImageJ software, as the percentage of the wound closed after 16 h of cell migration. Adhesion assays were carried out after transfection with Dbait32Hc or 8H by using a fibronectin-coated CytoSelect-48 cell adhesion assay (Cell Biolabs, San Diego, CA, USA) in accordance with the manufacturer's protocol. Briefly, cells were counted and plated (1610 5 cells in serum-free medium/well) on fibronectin-coated plates then incubated for 60 min at 37uC in a cell culture incubator. After incubation, adherent cells were stained and quantified at OD560nm after extraction.
We propose that beside the control of the immune response DNA-PK could play another anti-viral activity by modifying vimentin. In summary, we have shown that stabilized DSB-mimicking Dbait32Hc molecules may be used as a tool for studying DNA-PK-dependent signaling in cells. In this study, we demonstrated previously unknown target of DNA-PK which is IF vimentin. We mapped the DNA-PK phosphorylation sites of vimentin and confirmed the DNA-PK-dependent phosphorylation of vimentin at Ser459 in cells. We showed that Ser459-P-vimentin was mostly located at cell protrusions of migratory cells. We then demonstrated that vimentin phosphorylation at Ser459 induced by Dbait32Hc treatment leads to inhibition of cell adhesion and migration. Dbait32Hc treatment is sensitizing tumors to radiotherapy and is currently under clinical investigation for local metastatic melanoma treatment (clinicaltrials.
gov/show/ NCT01469455). Therefore, our results are very important as they provide evidence that Dbait32Hc treatment by inhibition of cancer cell migration can have anti-metastatic effect. Moreover, our results highlight the links between DNA-PK signaling, the cytoskeleton, cell adhesion and migration. These findings provide new insight into very important cellular processes controlled by DNA-PK. Table  S1  Detailed MS data including the Mascot Scores of the proteins identified in the 2D gel experiment. (PDF).
This work was supported by  Institut Curie ,  CNRS ,  INSERM , and the  Agence Nationale de la Recherche  grant  ANR-08-Biot-009-02  and  DNA Therapeutics SA. E. Kotula  and  M. Quanz  were recipients of a Ph. D. fellowship cofinanced by  DNA Therapeutics SA  and the  Agence Nationale de la Recherche ANRT. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Kotula is a recipient of a PhD fellowship cofinanced by  DNA Therapeutics  and  M. Quanz  is an employee of DNA Therapeutics. M.
A. Implementation of control and detection schemes The first step in the process of implementation of the control and detection units on a single FPGA chip is the fundamental simulation step. In this step, the studied system is modeled and simulated in the Matlab/Simulink environment, using Simulink and SimPowerSystem Toolbox. Initially continuous-time simulations are performed. Time-step in the simulations is variable and the model is in continuous time. System parameters are chosen to be equal to the parameters of the experimental setup, which is presented in the following. After validation, the control and detection blocks are discretized if necessary. In this case, the time-step is fixed and must be chosen small enough to guaranty a suitable accuracy. In this study it is chosen equal to 1 ‫.
ݏߤ‬ When the simulation results are validated, it is possible to proceed with mixed simulations. In the mixed simulation stage, the control blocks are replaced with proper DSP Builder blocks. DSP Builder provides the possibility of visual programming of a digital system in MATLAB-Simulink environment. Each DSP Builder block represents a VHDL module in the Simulink environment. This set of blocks is very efficient for rapid prototyping of FPGA devices. However some of the desired functions are not available in DSP Builder library and must be constructed from basic blocks or imported as form of HDL programming. The power part (fault tolerant converter, sources and charges) is remained unchanged in this step, modeled by using SimPowerSystem blocks. Appropriate DSP Builder input and output blocks are necessary to convert the Simulink signals to fixed point signals for DSP Builder.
B. HIL results For HIL experimentation, the controller and detection schemes are implemented on a single FPGA chip and the power system is modeled in MATLAB/Simulink environment using the SimPowerSystem toolbox. Here, an inverter application of the three-phase fault tolerant converter is investigated. A RL type load is placed at the AC side of the converter and the DC side is connected to a DC source with a voltage regulated at 300 V. A 2200 ‫ܨߤ‬ capacitance is used in the DC-link. Switching frequency is equal to 8 kHz. The time step and the sampling period are equal to 1 ‫. ݏߤ‬ N (see Fig. 4 ) is chosen equal to 30, to provide a fast, yet very robust fault detection. First, an open switch fault is applied to the upper switch of the leg 2 at t=0. 072s. Fig. 11  shows the three-phase  AC currents before and after the fault occurrence, fault detection and compensation.
Image processing was performed using the MRC suite of programs  (29). The phases were corrected for the effect of the CTF as calculated by determining the position of the first zero of the CTF (CTFFIND2). The crystals were tetragonal p42 1 2. The phase origins of the set of 18 selected best crystals were refined at successively higher resolution out to 4 A ˚by minimizing the average phase residual (ORIGTILTK). Defocus and astigmatism were refined for each image by comparison with the averaged phases from all the other images by using the program CTFSEARCH. We used an additional step of unbending (synthetic unbending), which notably improved the quality of the final map. In this supplementary unbending, the merged projection of the best single images is used as reference and each crystal is unbent against this reference.
The image amplitudes were scaled as a function of resolution to compensate for resolution-dependent attenuation by using the program SCALIMAM3D with bacteriorhodopsin as a reference. The phases were constrained to either 0 or 180 because the projection is centrosymmetric in this plane group. Amplitudes and phases from the images were averaged as described previously  (21). For some of the steps, we used the 2dx software  (30, 31).
3 A ). However, on images taken from samples tilted at high tilt angles (between 40 and 60 ), it was possible to count up to 10 layers of crystals, with a total thickness of ~0. 1 mm (Fig. 3 B ). Image analysis from 12 NS stacked crystals was done and results are shown in Fig. S2. The reason why the 2D crystals form stacks became clear when we solved the 3D structure of KirBac3. 1 using x-ray crystallography  (24). The analysis of the packing of the 3D crystals shows that these 3D crystals appear to be piles of 2D sheets, although no membrane surrounding the protein could be detected (Fig. S3 ). The absence of lipid membrane in these crystals was verified using the analysis of low-resolution diffraction intensities (data not shown, R. De Zorzi). This method has been reported previously for membrane proteins forming crystals of the so-called type I, i.
Morphological and Histological Evaluation Aorta segments were rapidly cleaned from the surrounding tissues and blood, cut into rings fixed in formalin 37% and embedded in paraffin blocks. Afterwards, five micrometer thick sections were cut with a rotational microtome (Leitz 1512, IMEB INC), placed onto glass microscope slides and stained with picrosirius red or hematoxylin and eosin by routine methods. Images from transverse sections of the arterial segments were captured using a camera connected to an optical microscope. Media and lumen areas were measured in triplicate and were quantified by planimetry using an image analyzer (LEICA Q550 IWB). In order to avoid miscalculation of the data due to possible deformation of the vessels during their preparation, we have determined luminal or vessel area by correcting the cross-sectional area enclosed by the internal or external elastic lamina, respectively, to a circle by applying the form factor l2/4P to the measurement of the lamina, where l is the length of the lamina.
Real Time Reverse-transcription PCR Total RNA was extracted with Trizol Reagent (Euromedex) and purified using the RNeasy kit, according to the manufacturer's instructions (Qiagen). First strand cDNA was synthesized according to the manufacturer's instructions (Roche). Quantitative PCR analysis was then performed with SYBR green PCR technology (ABGene) (Table  1 ). Relative quantification was achieved with MyiQ (Bio-rad) software according to the manufacturer's instructions. Data were normalized by HPRT levels and expressed as percentage relative to controls. All PCRs were performed at least in triplicate for each experimental condition.
Statistical Analysis Data are expressed as mean6SEM. Normality of distributions was verified by means of the Kolmogorov-Smirnov test. Pearson correlation analysis was used to examine association among different variables. Body weight and blood pressure were analyzed using a two-way analysis of variance, followed by a Bonferroni test. The rest of the data were analyzed using an unpaired t test, using GraphPad Software Inc. (San Diego, CA, USA). The predetermined significance level was a = 0. 05.
Electron Transfer activity Stopped-flow experiments were carried out with a thermostated instrument (DX. 17 MV, Applied Photophysics, Leatherhead, UK), equipped with a 1-cm pathlength observation chamber. Reactions were investigated by monitoring the absorption changes at selected wavelengths. When necessary, ionic strength was adjusted by addition of KCl and the buffer was degassed with vacuum/N 2 cycles. Data were analyzed using the software MATLAB (The Mathworks, South Natick, MA). Reduction by SOR TTHB8 of the cytochrome c 550 and its domains was investigated anaerobically at 45uC in 100 mM Tris-HCl pH 8. 0 buffer containing 0. 1 mM EDTA. To prevent inhibition of SOR TTHB8 resulting from prolonged incubation of the enzyme with a large excess of sulfite, in these experiments the stopped-flow instrument was used in the sequential mixing mode.
thermophilus HB8  [16]. Based on the high similarity between c 550 [N] and the SorB subunit of the sulfite:cytochrome c oxidoreductase from C. jejuni, it is very likely that this domain acts as an electron acceptor for SOR TTHB8. The 3D model of SOR TTHB8 was automatically built by means of the SWISS-MODEL server  [32] [33] [34] , using as a template the only available structure for a microbial SOR, i. e. , the one of the S. novella enzyme (PDB ID: 2c9x, segment A; 33% identity to SOR TTHB8 ) (Figure  3A ). In S. novella SOR, the formation of complementary electrostatic surfaces at the interface of the SorA and SorB subunit of sulfite:cytochrome c oxidoreductase has been observed  [35]. Similarly, a positively charged surface area surrounding the pocket cradling the molybdopterin cofactor is present also in the SOR from T.
Interestingly, the reaction between c 550 [N] and c 552 displays a similar ionic strength dependence to the full-length protein, whereas the dependence is less pronounced in the case of c 550 [C] (Figure  5C ). This suggests that electrostatic forces play an important role in the interaction between c 550 [N] and c 552 , while apolar interactions may be involved in molecular recognition between c 550 [C] and c 552. In order to test this possibility, the 3D model of c 550 [C] was automatically built by means of the SWISS-MODEL server  [32] [33] [34] , using the X-ray structure of cytochrome c 552 from T. thermophilus HB8 (PDB ID: 1c52; ,50% identity) as template (Figure  3BC ). The heme cleft in the very well described cytochrome c 552 is surrounded by a hydrophobic belt (Figure  3C ) consisting of residues G13, C14, F26, V68, M69 and F72; this patch of residues likely participates in the interaction between the cytochrome and the terminal oxidase ba 3  [36].
fruticosa Pall. ), a G2 QTL was identified in a similar linkage group position  (Wang et al. 2000). The CNR gene family provides an excellent source of candidate genes for investigating the genetic control of fruit size in Prunus. Its critical role of controlling fruit size by increasing cell number and organ size is demonstrated by FW2. 2 in tomato  (Frary et al. 2000) , and ZmCNR1 and ZmCNR2 in maize  (Guo et al. 2010). Using the peach genome v. 1. 0 sequence released by the International Peach Genome Initiative (GDR database: http://www. rosaceae. org/species/prunus_persica/gen ome_v1. 0), the identification of the CNR family in Prunus is possible. Because of the high level of synteny between peach and the other Prunus species  (Dirlewanger et al. 2004; Cabrera et al. 2009; Jung et al. 2009; Illa et al.