dataesr/scientific-paragraphs-categorization huggingface dataset:
Phylogenetic Reconstruction Sequence alignments were initially performed in MUSCLE  [25]  individually for each locus. Alignments were visually inspected and ambiguous regions were excluded in Mesquite 2. 5  [26]. Best-fit nucleotide substitution models were chosen through the Akaike information criterion, penalizing more complex models by one likelihood unit per additional free parameter, and ML phylogenetic trees for individual loci were estimated under these models in PAUP*  [27]. Conflict among the four loci was assessed through strong incongruence of nodes based on 1000 ML bootstrap replicates (. 70%) and posterior probabilities (. 99%) of credible Bayesian trees. Because no strongly supported nodes were in conflict, the data sets were combined into a single matrix with four partitions.
The partitions included 40, 154, 77, 728, 635, and 735 characters for ITS1, 5. 8 S, ITS2, LSU, EF1a and RPB2, respectively, for a combined data matrix of 2374 characters. Phylogenetic inferences were also conducted on a matrix consisting of 8 unlinked partitions: 1) SYM+I+G for 5. 8S and RPB2 position 1; 2) JC for RPB position 2; 3) K80+I+G for RPB position 3; 4) F81+I+G for EF1a position 1; 5) JC+I+G for EF1a position 2; 6) GTR+G for EF1a position 3; 7) SYM+G for ITS1 and ITS2; 8) K80+I+G for LSU. Parallel runs with four chains were allowed to run 20,000,000 generations, sampling every 1000 generations.
Divergence Time Estimation Molecular divergence time analyses were performed with the BEAST v1. 7. 2 software package  [30]  based on an alignment containing the four gene regions (ITS, LSU, EF1a, and RPB2) for a subset of the samples (one unique specimen per species -see above). Temporal calibration of divergence time analyses was achieved by fixing the absolute rate of molecular evolution for LSU locus (6. 5610 24 substitutions per site per million years)  [31]. The evolutionary rates of the ITS, EF1a and RPB2 regions were estimated relative to the fixed LSU rate using a relaxed clock model with an uncorrelated exponential prior distribution with a mean of 1. 0610 23 substitutions per site per million years assigned to the mean rate of each region. It is important to emphasize that this prior is on the mean of the rate of each locus, and rate heterogeneity is modeled at each of these loci by an exponential distribution to avoid over constraining the rate and rate variation at these loci.
The substitution models for the partitions were either HKY (RPB2) or GTR+G+I (LSU, ITS1, 5. 8S, ITS2, EF1a) substitution models. We used gene partitions rather than codon positions because when data partitions become small the ability to estimate parameters for the substitution model or clock models suffers. The birth/death speciation model was employed, and a fully resolved starting tree was provided for each analysis. Three independent and identical BEAST analyses were each run for 30 million generations, sampling parameters and trees every 1000 generations. Parameters from the resulting 30 thousand generations for each of the three runs were examined for convergence, stationarity, and suitable effective sample sizes in the program Tracer v1. 5  [33]. Based on this, a burn-in of 3000 trees was removed from each run, leaving 27,000 trees from each run, which were combined (81,000 trees) and used to generate a maximum clade credibility tree annotated with various parameter summary statistics using the program TreeAnnotator v1.
Ancestral Character State Reconstructions We reconstructed ancestral character states for EcM host plants of Tuber by phylum using the maximum likelihood model Markov k-state 1 parameter model in Mesquite  [26]. Hosts were coded either as gymnosperms, angiosperms, or both (Table  S1 ), considering a global database of Tuber ITS sequences that included host information were considered  [6]. Ancestral state reconstructions were also carried out on fruitbody type (epigeous vs. hypogeous) in the Tuberaceae and/gymnohydnotrya using an asymmetrical 2-parameter Markov-K model. In this model, parameter values for the transition from an epigeous to hypogeous fruiting body were relatively high ($106) compared to the transition from hypogeous to epigeous fruiting habit, reflecting the reality that in nature forcible spore discharge is more easily lost than reacquired  [21].
The program RASP  [34]  was used to statistically assess patterns of vicariance and dispersal across the genus using a distribution of equally probable Bayes trees and coding species by their geographical origins. Although Tuber aestivum, T. excavatum, T. puberulum, T. oligospermum and T. rufum have been reported from Northern Africa  [20] , collections from Africa were not available for study, and consequently, we did not include this biogeographic region in our analyses.
Phylogenetic Analyses Individual loci (Fig. 3 ) and combined molecular data (Fig. 4 ) confirm that the Tuberaceae is monophyletic as are both of the Northern hemisphere genera Tuber and Choiromyces. In contrast, genera of Southern hemisphere Tuberaceae were not resolved as monophyletic and are in need of taxonomic revision. Phylogenies of Tuber based on individual loci reconstructed the same major clades (Fig. 3 ). However, the LSU phylogeny does not resolve the/ puberulum lineage or place T. magnatum within the/aestivum lineage. Because there was no strongly supported conflict between single gene phylogenies, we combined the data sets to improve phylogenetic resolution. Eleven major clades can be recognized within Tuber based on the concatenated dataset (Figs. 4  & 5 ). The/ rufum,/melanosporum,/puberulum,/maculatum, and/macrosporum clades are distributed across the entire Northern hemisphere (Europe, Asia, North America, Central America and Northern Africa), yet are characterized by a high degree of specieslevel endemism.
For instance, Kinoshita et al. [22]  recently reported more than 20 undescribed Tuber species in Japan, including members of the/japonicum lineage. The addition of phylogenetically dispersed representatives of these species provided insights on the historical biogeography of Tuber, but the relationship between the/japonicum and/gennadii lineages to the rest of Tuber was still not fully resolved with this dataset. While we must reject a strict vicariance model for explaining the biogeography of Tuber, particularly for the/puberulum group that is the most widely distributed clade, most species and many Tuber clades appear to have restricted distributions at the continental scale in accordance with our hypothesis. For instance, the/ gibbosum,/japonicum,/gennadii, and/multimaculatum clades appear to be restricted to single continents.
Acknowledgments Many thanks to the North American Truffling Society and other collectors who provided the material used in this research. Particularly,  Joey Spatafora  and  Richard Halse  of the  Oregon State University Department of Botany Herbarium  for extensive herbarium services for accessions of the collections cited here plus many additional collections. Connie Robertson  for curating truffle collections at the  Duke Herbarium. The  National Fungus Collections and Herbarium of the Universitadi Bologna  generously lent specimens for study. We thank  Paul Manos ,  David Swofford ,  Michael Castellano ,  Hiromi Sasaki ,  Pei-Gui Liu ,  Yongjin Wang ,  Juan Chen ,  Francois Le Tacon ,  Terri Porter ,  Jason Jackson ,  Andrii Gryganskyi  and  Michelle Hersh  for providing specimens, sequences, photos or valuable discussion for the preparation of this manuscript.
Unlinking measurements were performed with 5 nM topo III in topo III activity buffer [40 mM HEPES, pH 8. 0 adjusted with KOH, 1 mM magnesium acetate, pH 7, 0. 3% (w/v) BSA and 0. 04% (v/v) Tween-20]. Data traces were analysed using a custom step-finding algorithm to identify the waiting time, T wait (the time interval between rotating the bead to generate the DNA link and it being unlinked by topoisomerase). The T wait values (100-200 events for each rotation) were binned, and the histogram was fitted with a single exponential to obtain the strand-passage rate. Topo I (New England Biolabs) unlinking measurements were performed using the same substrate used for topo III experiments with $130 or 325 nM of topo I in topo I activity buffer [50 mM potassium acetate, pH 7. 9, 10 mM magnesium acetate, 1 mM dithiothreitol, 0.
ACKNOWLEDGEMENTS The authors thank  James Keck  for providing the topo III expression construct,  Richard Neuman  and  Tamara Litwin  for editing the manuscript,  Krishan Zaveri  for assistance with the gapped template construction,  Alfonso Mondrago´n  for useful discussions and  David Bensimon  and  Vincent Croquette  for supporting and encouraging this work in its initial phase. FUNDING Intramural Research Program  of the  National Heart, Lung, and Blood Institute, National Institutes of Health  (in part);  Young Investigator Award  from the  Human Frontiers Science Program  (to K. C. N. ) (in part). Funding for open access charge: The  Intramural Research Program  of the  National Heart, Lung, and Blood Institute, National Institutes of Health.
Back Close Full The dam inflows are measured daily since 1984 at the Makhazine station. The daily pan evaporation measured between 1984 and 2011 and monthly air temperatures between 1975 and 1996 are also available. 11 stations with daily precipitation (P ) located within and outside the catchment were considered (Fig. 1 ). The study period is 1984-2010 (27 hydrological years from September to August). The rainfall depth at the catchment scale was obtained through spatial interpolation with the Thiessen approach. No trends were detected in precipitation or temperature data aggregated over annual or monthly time steps between 1984 and 2010. The individual precipitation records at each station have been carefully checked to remove unrealistic values and identify the missing days/periods.
, 2010). Different configurations of this model can be used. They are obtained by varying the spatial resolution, the size and position of the area of interest, and the dataset used for the large scale forcing. More complex and unusual options are also available such as the use of spectral nudging and interactive air-sea coupling techniques (see  Herrmann et al. , 2011). Those options are not used in the current study. For the model definition, we used a Lambert conformal projection for pan-Mediterranean area at horizontal resolutions of 50 km (resp. 12 km) centred at 14 • E, 43 • N with 128 (resp. 432) grid points in longitude and 90 (resp. 288) grid points in latitude including the bi-periodization (11) and the relaxation zone (2 × 8). The model version has 31 vertical levels. The time step used is 1800 s.
Quantile perturbation A quantile perturbation method does not consider the differences between observed and simulated datasets but only the relative changes between past and future time periods as simulated by climate models. Several authors have implemented this method for hydrological impact studies  (Chiew et al. , 2009; Liu et al. , 2011; Willems and Vrac, 2011) : we introduced a variant that is producing random ensembles of perturbed series, to prevent that a single random generation may contain outliers. The computation was done for each month separately, between an observed period (CTRL), a simulation during the observation period (SIM) and a projected future period (PROJ). The approach followed these steps: 1. Identification of the frequency of dry days (when P < 0. 1 mm) in CTRL, SIM and PROJ; 2.
, 2012; Milano et al. , 2012 Milano et al. , , 2013)). Further work should now test other approaches to evaluate the climate change signal and compare the results with those obtained in the present study. In particular other types of bias-correction methods and hydrological models should be considered but also the use of RCM with an improved description of the present-climate hydrological cycle (improved mean behavior and extreme events). We are however aware that individual RCM simulations do not allow to take into account two of the main sources of uncertainty in regional climate change, that is to say the choice of the RCM and the choice of the forcing GCM  (D équ é et al. , 2012). This will be possible in the future with the extension of the number of Med-CORDEX simulations and database (www.
HESSD HESSD Acknowledgements. The observed data was provided by the  department of Water Research and Planning (DRPE) of Morocco  and the  Hydraulic Basin Agency of Loukkos-T étouan. We thank  A. Alias  ( CNRM ) for running the Mediterranean ALADIN simulations. This work is a contribution to the  HyMeX program  (HYdrological cycle in the Mediterranean Experiment) through INSU-MISTRALS support and to the  Med-CORDEX initiative (COordinated Regional climate Downscaling EXperiment -Mediterranean region ). The third author has received funding from the  French National Research Agency (ANR)  project  REMEMBER  (contract  ANR-12-SENV-001 ) and from the  European Commission  through the  FP7   CLIM-RUN  project. The ALADIN simulations used in the current work can be downloaded from the Med-CORDEX database (www.
ACKNOWLEDGEMENTS We thank  Dr G. Felsenfeld   Institute of Diabetes and Digestive and Kidney Diseases, USA)  for the HS4 insulator sequences. FUNDING  Defence Advanced Research Projects Agency  [ HR0011-12-C-0067 ,  DARPA-BAA-11-23 ];  National Science Foundation  [ CBET-0939511 ];  Synthetic Biology Engineering Research Center (SynBERC)  [ SA5284-11210 ];  National Institutes of Health  [ 5-R01-CA155320-02 ];  Swiss National Science Foundation  [ PA00P3_131492  to P. G. ]; the  Ernst Schering Foundation  (to P. G. ). Funding for open access charge:  SynBERC  [ SA5284-11210 ].
The 2011 harvest was used for analysis by 454 to improve the sequence assembly. Desiccation sensitivity of mature C. australe cotyledons was determined on 3 × 5 × 3 mm cubes that were isolated from the inner part of the cotyledons. Cubes were dried for the indicated time intervals over a saturated salt solution at 75% RH, after which they were divided into two halves. One half was used for water content determination, and the other half for viability assessment following incubation for 24 h in a 1% (w/v) tetrazolium solution (Sigma-Aldrich, France). Red colour was quantified by pixel intensity on the image using ImageJ software (http://rsb. info. nih. gov/ ij/). Water content was determined gravimetrically by weighing the seeds before and after drying in an oven for 48 h at 96 °C. Viability assays were performed on four independent drying experiments of 50-100 cubes.
Cloning of MtABI3 To obtain the full-length sequence for MtABI3, genomic DNA was extracted from leaf material of M. truncatula A17 using the Nucleospin Food kit (Macherey Nagel). An inverse PCR was performed on 5 μg of genomic DNA that was digested with EcoRI (25 U per 50 μl final volume; Promega, Madison, WI, USA), and ligated using T4 DNA ligase (50 U per 450 μl final volume; Fermentas, Vilnius, Lithuania). The full length was amplified on the ligated DNA using the primers indicated in Supplementary Table  S1  at JXB online that were designed based on the MtABI3 fragment available in the expresssed sequence tag (EST) database (TC97588, the DFCI Medicago truncatula Gene Index v8). The full-length genomic DNA fragment (3 458 bp) was cloned into pJet1. 2 (CloneJET kit, Thermo Scientific, Bremen, Germany) and sequenced (for primers see Supplementary Table  S1 ).
From the 2011 RNA pool, a cDNA library was prepared, normalized, and sequenced by Eurofins (Ebersberg, Germany) using the 454 GS FLX+ technology. Reads obtained from each sequencing were assembled de novo in two steps: first with MIRA 3. 4. 0 (Chevreux et al. , 2004)  then with DNA Dragon (SequentiX, http://www. sequentix. de/software_dnadragon. php). The detailed procedure is described in Supplementary Fig. S1  at JXB online.
Functional annotation and classification Contig annotation to known sequences by sequence similarity was performed using two M. truncatula nucleic databases: MT3. 5 from the International Medicago Genome Annotation Group (IMGAG) and MtGI11 from the Dana-Farber Cancer Institute (DFCI) Medicago Gene Index. Contigs that remained unannotated after these two analyses were blasted using Blast2GO (version 2. 6. 0) (Götz et al. , 2008)  against protein databases including all plant species: Swissprot and non-redundant protein from NCBI. Next, classification of C. australe annotations in Gene Ontology (GO) was performed by Blast2GO. GO terms were retrieved from public databases and mapped to each contig, after which the most specific ones were selected by an annotation rule. The detailed annotation workflow is described in Supplementary Fig.
(2006). Mass data were analysed with the Protein Lynx Global Server software (Micromass-Waters). Protein identification was performed by comparing the data with the UniProt sequence databank (date of release: August 2010) or with the TIGR Medicago EST databank (date of release: April 2010). For the M. truncatula heat-stable proteome, spots linked to LEA polypeptides were identified according to the reference gel published by  Chatelain et al. (2012).
Data submission Raw sequence data from this article can be found in the Sequence Reads Archive database (NCBI) under BioProject PRJNA193308. The data on the ectopic expression of MtABI3 in hairy roots discussed in this publication have been deposited in NCBI's Gene Expression Omnibus  (Edgar et al. , 2002)  and are accessible through GEO Series accession number GSE44291 (http://www. ncbi. nlm. nih. gov/geo/query/acc. cgi?acc=GSE44291).
5 g H 2 O g DW -1 was reached, after which the intensity decreased progressively with further drying (Fig. 1E ). The critical water content, here defined as the water content corresponding to the break point for which cotyledon tissues begin to lose staining intensity, was estimated at 1. 2 g H 2 O g DW -1. Tissues were completely lacking red staining when dried below 0. 5 g H 2 O g DW -1 (Fig. 1D ), indicating total loss of viability. This value is consistent with that reported by  Han et al. (1997)  on isolated axes of C. australe during rapid drying using electrolyte conductivity as an indication of membrane damage. In contrast, complete drying and rehydration of 10 h imbibed M. truncatula cotyledons in tetrazolium solution rendered the tissues red, indicating that viability was maintained (data not shown).
S1  at JXB online). A total of 35 050 contigs (72. 5%) were annotated, of which 91% were provided by the IMGAG 3. 5 M. truncatula database and the MtGI11 version of the Medicago EST database. An additional 740 and 2558 of the remaining contigs were identified, respectively, using Swissprot and NR databases related to other plant species (Supplementary Fig. S1 ). Annotations were classified according to GO using the Plant-GO-slim version of Blast2GO (B2G). Enzyme classification (EC) numbers were retrieved with the additional functionalities of B2G linked to KEGG pathways. A total of 23 637 contigs (48. 9%) were annotated with 98 615 GO terms and 8 962 EC numbers. The distribution of the main biological processes (BP, 45 568 annotations), molecular functions (MF, 35 110 annotations), and cellular components (CC, 17 937 annotations) is shown in Supplementary Fig.
truncatula (GeOmnibus GS GSE44291). The advantage of this ectopic expression model is that it avoids the interfering effects of other B3 domain transcription factors such as FUS3 and LEC2 with ABI3 in seeds  (Mönke et al. , 2012). Transcript levels of 9 out of 11 genes coding for group A LEA proteins were up-regulated by MtABI3 (Table  3 ). Moreover, in silico promoter analysis of 8 out of 10 LEA genes for which promoter sequences could be retrieved indicated that seven LEA promoters from group A contain both RY (CATGCA) and ABRE (ACGTG(G/T)C) cis-regulatory elements. The RY element is known to be bound by ABI3, while ABRE motifs are implicated in the binding of bZIP-TFs, known to interact with ABI3  (Busk et al. , 1997; Hattori et al. , 2002; Guerriero et al. , 2009). A study on the identification of the ABI3 regulon in Arabidopsis confirmed three LEA genes as direct targets by transient promoter activation assay or ChIP-chip analysis (Table  3 ).
(2011). More than 72% of the 48 334 contigs could be annotated by this approach, and 91% of this annotation is provided by M. truncatulaspecific databases. This approach also enabled the discovery of almost all LEA transcripts for comparison with M. truncatula. However, a quantitative transcriptome analysis will be needed to reveal to what extent LEA polypeptide abundance is regulated at the transcriptional and/or post-transcriptional level in C. australe. Furthermore, there are many other molecular protective mechanisms that could be missing in this recalcitrant species such as antioxidant defences, nonreducing sugars, heat shock proteins, and/or induction of cell wall modifications (reviewed in  Berjak and Pammenter, 2008; Leprince and Buitink, 2010). The sequence assembly from the normalized library will enable the construction of microarrays to investigate further molecular aspects of desiccation sensitivity in recalcitrant seeds.
Acknowledgements We thank our South-African colleagues ( Professor P. Berjak ,  Professor N. Pammenter , and  Professor J. Farrant ) for help with collecting the material. The abi3 mutants of Medicago truncatula utilized in this research project, which are jointly owned by the  Centre National de la Recherche Scientifique , were obtained from The  Samuel Roberts Noble Foundation, Inc. and were created through research funded, in part, by a grant from the  National Science Foundation  (NSF#  703285 ). No conflict of interest is declared. This work was supported by a grant from the  Région des Pays-de-la-Loire, France  ( QUALISEM 2009 ( QUALISEM -2013 )), the  bilateral Partenariat Hubert Curien (PHC) program France-South Africa  (grant no. 25903RE  to JD, OL, and JB), and a post-doctoral  EU   FP7 Marie Curie Individual Fellowship  (grant no.
1 9 LEAm Mt3. 5 Medtr2g014040 Ca_7604 3. E-45 268 48. 6 62. 4 74 CAPLEA. I MtGI11 TC175990 Ca_8841 1. E-31 141 66. 0 85. 8 18,19 LEA_1 PM1 Mt3. 5 Medtr7g093170 Ca_8304 3. E-16 91 64. 8 75. 8 ND D113. II Mt3. 5 Medtr7g093160 Ca_8304 5. E-17 91 65. 2 76. 1 ND SMP PM25 MtGI11 TC174777 Ca_6007 2. E-83 238 75. 6 84. 4 92 D-34. I MtGI11 Medtr1g072090 ND ND D-34. II MtGI11 TC183570 Ca_25629 6. E-42 128 86. 7 93 ND D-34. III Mt3. 5 Medtr2g076230 Ca_23377 3. E-06 24 76. 0 87. 5 ND ND, homologue not detected. Contigs were translated to calculate the percentage identity and similarity. a Castanospermum australe contigs that were homologues to the 17 LEA gene products detected in M. truncatula seeds were identified based on the MtGI11 and Mt3. 5 Medicago databases (E-value <e-06).
06 6. 96E-02 ND AT1G52690 Up - Mönke et al. (2012)(2008) levels(and corresponding P-values)in hairy roots overexpressing MtABI3 compared with control (empty plasmid), determined by trancriptome analysis using Nimblegen slides (GeOmnibus GSE44291). bThenumber of RY (CATGCA) and ABRE (ACGTG(G/T)C) cis-regulatory motifs known to bind ABI3 was revealed by analysing the 2 kb promoter sequence of the M. truncatula genes. cRelativelevel of transcripts in mature abi3 seeds of Arabidopsis compared with the wild type. Data are extracted fromBies-Etheve et al. (2008). dIdentification of ABI3-responsive gene products in 35S::ABI3-GR seedlings by array-based transcriptome analysis (T) or qRT-PCR (P) and confirmation as direct targets by transient promoter activation assay (A) or ChIP-chip analysis (C).