Trichoderma harzianum expressed sequence tags for identification of genes with putative roles in mycoparasitism against Fusarium solani

The plant pathogen Fusarium solani causes a disease root rot of common bean (Phaseolus vulgaris) resulting in great losses of yield in irrigated areas of the Southeast and Midwest regions of Brazil. Species of the genus Trichoderma have been used in the biological control of this pathogen as an alternative to chemical control. To gain new insights into the biocontrol mechanism used by Trichoderma harzianum against the phytopathogenic fungus, Fusarium solani, we performed a transcriptome analysis using expressed sequence tags (ESTs) and quantitative real-time PCR (RT-qPCR) approaches. A cDNA library from T. harzianum mycelium (isolate ALL42) grown on cell walls of F. solani (CWFS) was constructed and analyzed. A total of 2927 high quality sequences were selected from 3845 and 37.7% were identified as unique genes. The Gene Ontology analysis revealed that the majority of the annotated genes are involved in metabolic processes (80.9%), followed by cellular process (73.7%). We tested twenty genes that encode proteins with potential role in biological control. RT-qPCR analysis showed that none of these genes were expressed when T. harzianum was challenged with itself. These genes showed different patterns of expression during in vitro interaction between T. harzianum and F. solani. (C) 2012 Elsevier Inc. All rights reserved.

SIS: a program to generate draft genome sequence scaffolds for prokaryotes

Background: Decreasing costs of DNA sequencing have made prokaryotic draft genome sequences increasingly common. A contig scaffold is an ordering of contigs in the correct orientation. A scaffold can help genome comparisons and guide gap closure efforts. One popular technique for obtaining contig scaffolds is to map contigs onto a reference genome. However, rearrangements that may exist between the query and reference genomes may result in incorrect scaffolds, if these rearrangements are not taken into account. Large-scale inversions are common rearrangement events in prokaryotic genomes. Even in draft genomes it is possible to detect the presence of inversions given sufficient sequencing coverage and a sufficiently close reference genome. Results: We present a linear-time algorithm that can generate a set of contig scaffolds for a draft genome sequence represented in contigs given a reference genome. The algorithm is aimed at prokaryotic genomes and relies on the presence of matching sequence patterns between the query and reference genomes that can be interpreted as the result of large-scale inversions; we call these patterns inversion signatures. Our algorithm is capable of correctly generating a scaffold if at least one member of every inversion signature pair is present in contigs and no inversion signatures have been overwritten in evolution. The algorithm is also capable of generating scaffolds in the presence of any kind of inversion, even though in this general case there is no guarantee that all scaffolds in the scaffold set will be correct. We compare the performance of SIS, the program that implements the algorithm, to seven other scaffold-generating programs. The results of our tests show that SIS has overall better performance. Conclusions: SIS is a new easy-to-use tool to generate contig scaffolds, available both as stand-alone and as a web server. The good performance of SIS in our tests adds evidence that large-scale inversions are widespread in prokaryotic genomes.

Probing the anomalous extinction of four young star clusters: the use of colour-excess, main-sequence fitting and fractal analysis

Aims. We studied four young star clusters to characterise their anomalous extinction or variable reddening and asses whether they could be due to contamination by either dense clouds or circumstellar effects. Methods. We evaluated the extinction law (R-V) by adopting two methods: (i) the use of theoretical expressions based on the colour-excess of stars with known spectral type; and (ii) the analysis of two-colour diagrams, where the slope of the observed colour distribution was compared to the normal distribution. An algorithm to reproduce the zero-age main-sequence (ZAMS) reddened colours was developed to derive the average visual extinction (A(V)) that provides the closest fit to the observational data. The structure of the clouds was evaluated by means of a statistical fractal analysis, designed to compare their geometric structure with the spatial distribution of the cluster members. Results. The cluster NGC 6530 is the only object of our sample affected by anomalous extinction. On average, the other clusters suffer normal extinction, but several of their members, mainly in NGC 2264, seem to have high R-V, probably because of circumstellar effects. The ZAMS fitting provides A(V) values that are in good agreement with those found in the literature. The fractal analysis shows that NGC 6530 has a centrally concentrated distribution of stars that differs from the substructures found in the density distribution of the cloud projected in the A(V) map, suggesting that the original cloud was changed by the cluster formation. However, the fractal dimension and statistical parameters of Berkeley 86, NGC 2244, and NGC 2264 indicate that there is a good cloud-cluster correlation, when compared to other works based on an artificial distribution of points.

Time sequence of the intensification of the liver glucose production induced by high-fat diet in mice

It is well established that the development of insulin resistance shows a temporal sequence in different organs and tissues. Moreover, considering that the main aspect of insulin resistance in liver is a process of glucose overproduction from gluconeogenesis, we investigated if this metabolic change also shows temporal sequence. For this purpose, a well-established experimental model of insulin resistance induced by high-fat diet (HFD) was used. The mice received HFD (HFD group) or standard diet (COG group) for 1, 7, 14 or 56?days. The HFD group showed increased (P?<?0.05 versus COG) epididymal, retroperitoneal and inguinal fat weight from days 1 to 56. In agreement with these results, the HFD group also showed higher body weight (P?<?0.05 versus COG) from days 7 to 56. Moreover, the changes induced by HFD on liver gluconeogenesis were progressive because the increment (P?<?0.05 versus COG) in glucose production from l-lactate, glycerol, l-alanine and l-glutamine occurred 7, 14, 56 and 56 days after the introduction of the HFD schedule, respectively. Furthermore, glycaemia and cholesterolemia increased (P?<?0.05 versus COG) 14?days after starting the HFD schedule. Taken together, the results suggest that the intensification of liver gluconeogenesis induced by an HFD is not a synchronous all-or-nothing process but is specific for each gluconeogenic substrate and is integrated in a temporal manner with the progressive augmentation of fasting glycaemia. Copyright (c) 2012 John Wiley & Sons, Ltd.

Identical sequence patterns in the ends of exons and introns of human protein-coding genes

Intron splicing is one of the most important steps involved in the maturation process of a pre-mRNA. Although the sequence profiles around the splice sites have been studied extensively, the levels of sequence identity between the exonic sequences preceding the donor sites and the intronic sequences preceding the acceptor sites has not been examined as thoroughly. In this study we investigated identity patterns between the last 15 nucleotides of the exonic sequence preceding the 5' splice site and the intronic sequence preceding the 3' splice site in a set of human protein-coding genes that do not exhibit intron retention. We found that almost 60% of consecutive exons and introns in human protein-coding genes share at least two identical nucleotides at their 3' ends and, on average, the sequence identity length is 2.47 nucleotides. Based on our findings we conclude that the 3' ends of exons and introns tend to have longer identical sequences within a gene than when being taken from different genes. Our results hold even if the pairs are non-consecutive in the transcription order. (C) 2012 Elsevier Ltd. All rights reserved.

Hero's journey in bifurcation diagram

The hero's journey is a narrative structure identified by several authors in comparative studies on folklore and mythology. This storytelling template presents the stages of inner metamorphosis undergone by the protagonist after being called to an adventure. In a simplified version, this journey is divided into three acts separated by two crucial moments. Here we propose a discrete-time dynamical system for representing the protagonist's evolution. The suffering along the journey is taken as the control parameter of this system. The bifurcation diagram exhibits stationary, periodic and chaotic behaviors. In this diagram, there are transition from fixed point to chaos and transition from limit cycle to fixed point. We found that the values of the control parameter corresponding to these two transitions are in quantitative agreement with the two critical moments of the three-act hero's journey identified in 10 movies appearing in the list of the 200 worldwide highest-grossing films. (C) 2011 Elsevier B.V. All rights reserved.

Sequence and Copy Number Analyses of HEXB Gene in Patients Affected by Sandhoff Disease: Functional Characterization of 9 Novel Sequence Variants

Sandhoff disease (SD) is a lysosomal disorder caused by mutations in the HEXB gene. To date, 43 mutations of HEXB have been described, including 3 large deletions. Here, we have characterized 14 unrelated SD patients and developed a Multiplex Ligation-dependent Probe Amplification (MLPA) assay to investigate the presence of large HEXB deletions. Overall, we identified 16 alleles, 9 of which were novel, including 4 sequence variation leading to aminoacid changes [c.626C>T (p.T209I), c.634C>A (p.H212N), c.926G>T (p.C309F), c.1451G>A (p.G484E)] 3 intronic mutations (c.1082+5G>A, c.1242+1G>A, c.1169+5G>A), 1 nonsense mutation c.146C>A (p.S49X) and 1 small in-frame deletion c.1260_1265delAGTTGA (p.V421_E422del). Using the new MLPA assay, 2 previously described deletions were identified. In vitro expression studies showed that proteins bearing aminoacid changes p.T209I and p.G484E presented a very low or absent activity, while proteins bearing the p.H212N and p.C309F changes retained a significant residual activity. The detrimental effect of the 3 novel intronic mutations on the HEXB mRNA processing was demonstrated using a minigene assay. Unprecedentedly, minigene studies revealed the presence of a novel alternative spliced HEXB mRNA variant also present in normal cells. In conclusion, we provided new insights into the molecular basis of SD and validated an MLPA assay for detecting large HEXB deletions.

Lithium in M 67: From the main sequence to the red giant branch

Context. Lithium abundances in open clusters are a very effective probe of mixing processes, and their study can help us to understand the large depletion of lithium that occurs in the Sun. Owing to its age and metallicity, the open cluster M 67 is especially interesting on this respect. Many studies of lithium abundances in M 67 have been performed, but a homogeneous global analysis of lithium in stars from subsolar masses and extending to the most massive members, has yet to be accomplished for a large sample based on high-quality spectra. Aims. We test our non-standard models, which were calibrated using the Sun with observational data. Methods. We collect literature data to analyze, for the first time in a homogeneous way, the non-local thermal equilibrium lithium abundances of all observed single stars in M 67 more massive than similar to 0.9 M-circle dot. Our grid of evolutionary models is computed assuming a non-standard mixing at metallicity [Fe/H] = 0.01, using the Toulouse-Geneva evolution code. Our analysis starts from the entrance into the zero-age main-sequence. Results. Lithium in M 67 is a tight function of mass for stars more massive than the Sun, apart from a few outliers. A plateau in lithium abundances is observed for turn-off stars. Both less massive (M >= 1.10 M-circle dot) and more massive (M >= 1.28 M-circle dot) stars are more depleted than those in the plateau. There is a significant scatter in lithium abundances for any given mass M <= 1.1 M-circle dot. Conclusions. Our models qualitatively reproduce most of the features described above, although the predicted depletion of lithium is 0.45 dex smaller than observed for masses in the plateau region, i.e. between 1.1 and 1.28 solar masses. More work is clearly needed to accurately reproduce the observations. Despite hints that chromospheric activity and rotation play a role in lithium depletion, no firm conclusion can be drawn with the presently available data.

Facial Profile Evaluation of Isolated Pierre Robin Sequence

Objective: To evaluate numerically the facial profile of children with isolated Pierre Robin sequence (PRS) and to compare them with a control group that has no pathologies and exhibits regular and balanced facial growth, with no skeletal alterations. Patients: Eighty-three children aged 5 to 10 years (PRS group, n = 60; control group, n = 23) were selected. Setting: Hospital for Rehabilitation of Craniofacial Anomalies, University of Sao Paulo (HRAC-USP). Children from the control group were taken from the program of Interceptive Orthodontics at HRAC-USP. Design: Angular and ratio analyses of the facial profiles in both groups were realized through digital photographs. The PRS group was subdivided into two groups-complete and incomplete-according to the sagittal extension of the cleft palate, to investigate the possible influence of cleft extension on the face. Results: The facial convexity angle and the facial inferior third angle were considerably higher in the PRS groups than in the control group and were not significantly different between PRS groups. Nasolabial angle did not differ between groups. Conclusion: The facial profile was more convex in individuals with PRS than in those with regular facial growth and with no pathology. The mandible was responsible for the convexity of the profile in PRS because of its lack off anterior projection. An important relationship between the extension of the cleft palate and alterations in facial profile in PRS was not observed.

Characterisation of young stellar clusters

Aims. Several embedded clusters are found in the Galaxy. Depending on the formation scenario, most of them can evolve to unbounded groups that are dissolved within 10 Myr to 20 Myr. A systematic study of young stellar clusters that show distinct characteristics provides interesting information on the evolutionary phases during the pre-main sequence. To identify and to understand these phases we performed a comparative study of 21 young stellar clusters. Methods. Near-infrared data from 2MASS were used to determine the structural and fundamental parameters based on surface stellar density maps, radial density profile, and colour-magnitude diagrams. The cluster members were selected according to their membership probability, which is based on the statistical comparison with the cluster proper motion. Additional members were selected on the basis of a decontamination procedure that was adopted to distinguish field stars found in the direction of the cluster area. Results. We obtained age and mass distributions by comparing pre-main sequence models with the position of cluster members in the colour-magnitude diagram. The mean age of our sample is similar to 5 Myr, where 57% of the objects is found in the 4-10 Myr range of age, while 43% is <4 Myr old. Their low E(B - V) indicate that the members are not suffering high extinction (AV <1 mag), which means they are more likely young stellar groups than embedded clusters. Relations between structural and fundamental parameters were used to verify differences and similarities that could be found among the clusters. The parameters of most of the objects show the same trends or correlations. Comparisons with other young clusters show similar relations among mass, radius, and density. Our sample tends to have larger radius and lower volumetric density than embedded clusters. These differences are compatible with the mean age of our sample, which we consider intermediate between the embedded and the exposed phases of the stellar clusters evolution.

Effect of sequence of insemination after simultaneous thawing of multiple semen straws on conception rate to timed AI in suckled multiparous Nelore cows

The objective was to determine the effect of sequence of insemination after simultaneous thawing of multiple 0.5 mL semen straws on conception rate in suckled multiparous Nelore cows. The effect of this thawing procedure on in vitro sperm characteristics was also evaluated. All cows (N = 944) received the same timed AI protocol. Ten straws (0.5 mL) of frozen semen from the same batch were simultaneously thawed at 36 degrees C, for a minimum of 30 sec. One straw per cow was used for timed AI. Frozen semen from three Angus bulls was used. Timed AI records included sequence of insemination (first to tenth) and time of semen removal from thawing bath. For laboratory analyses, the same semen batches used in the field experiment were evaluated. Ten frozen straws from the same batch were thawed simultaneously in a thawing unit identical to that used in the field experiment. The following sperm characteristics were analyzed: sperm motility parameters, sperm thermal resistance, plasma and acrosomal membrane integrity, lipid peroxidation, chromatin structure, and sperm morphometry. Based on logistic regression, there were no significant effects of breeding group, body condition score, AI technician, and sire on conception rate, but there was an interaction between sire and straw group (P = 0.002). Semen from only one bull had decreased (P < 0.05) field fertility for the group of straws associated with the longest interval from thawing to AI. However, the results of the laboratory experiment were unable to explain the findings of the field experiment. Sperm width:length ratio of morphometric analysis was the single sperm characteristic with a significant interaction between sire and straw group (P = 0.02). It was concluded that sequence of insemination after simultaneous thawing of 10 semen straws can differently affect conception rates at timed AI, depending on the sire used. Nevertheless, the effects of this thawing environment on in vitro sperm characteristics, remain to be further investigated. (C) 2012 Elsevier Inc. All rights reserved.

Use of Carr-Purcell pulse sequence with low refocusing flip angle to measure T-1 and T-2 in a single experiment

The Carr-Purcell pulse sequence, with low refocusing flip angle, produces echoes midway between refocusing pulses that decay to a minimum value dependent on T*(2). When the refocusing flip angle was pi/2 (CP90) and tau > T*(2), the signal after the minimum value, increased to reach a steady-state free precession regime (SSFP), composed of a free induction decay signal after each pulse and an echo, before the next pulse. When tau < T*(2), the signal increased from the minimum value to the steady-state regime with a time constant (T*) = 2T(1)T(2)/(T-1 + T-2). identical to the time constant observed in the SSFP sequence, known as the continuous wave free precession (CWFP). The steady-state amplitude obtained with M-cp90 = M0T2/(T-1+T-2) was identical to CWFP. Therefore, this sequence was named CP-CWFP because it is a Carr-Purcell sequence that produces results similar to the CWFP. However, CP-CWFP is a better sequence for measuring the longitudinal and transverse relaxation times in single scan, when the sample exhibits T-1 similar to T-2. Therefore, this sequence can be a useful method in time domain NMR and can be widely used in the agriculture, food and petrochemical industries because those samples tend to have similar relaxation times in low magnetic fields. (C) 2011 Elsevier Inc. All rights reserved.

A UML profile for the OBO relation ontology

Background: Ontologies have increasingly been used in the biomedical domain, which has prompted the emergence of different initiatives to facilitate their development and integration. The Open Biological and Biomedical Ontologies (OBO) Foundry consortium provides a repository of life-science ontologies, which are developed according to a set of shared principles. This consortium has developed an ontology called OBO Relation Ontology aiming at standardizing the different types of biological entity classes and associated relationships. Since ontologies are primarily intended to be used by humans, the use of graphical notations for ontology development facilitates the capture, comprehension and communication of knowledge between its users. However, OBO Foundry ontologies are captured and represented basically using text-based notations. The Unified Modeling Language (UML) provides a standard and widely-used graphical notation for modeling computer systems. UML provides a well-defined set of modeling elements, which can be extended using a built-in extension mechanism named Profile. Thus, this work aims at developing a UML profile for the OBO Relation Ontology to provide a domain-specific set of modeling elements that can be used to create standard UML-based ontologies in the biomedical domain. Results: We have studied the OBO Relation Ontology, the UML metamodel and the UML profiling mechanism. Based on these studies, we have proposed an extension to the UML metamodel in conformance with the OBO Relation Ontology and we have defined a profile that implements the extended metamodel. Finally, we have applied the proposed UML profile in the development of a number of fragments from different ontologies. Particularly, we have considered the Gene Ontology (GO), the PRotein Ontology (PRO) and the Xenopus Anatomy and Development Ontology (XAO). Conclusions: The use of an established and well-known graphical language in the development of biomedical ontologies provides a more intuitive form of capturing and representing knowledge than using only text-based notations. The use of the profile requires the domain expert to reason about the underlying semantics of the concepts and relationships being modeled, which helps preventing the introduction of inconsistencies in an ontology under development and facilitates the identification and correction of errors in an already defined ontology.

Decreasing the number of false positives in sequence classification

Abstract Background A large number of probabilistic models used in sequence analysis assign non-zero probability values to most input sequences. To decide when a given probability is sufficient the most common way is bayesian binary classification, where the probability of the model characterizing the sequence family of interest is compared to that of an alternative probability model. We can use as alternative model a null model. This is the scoring technique used by sequence analysis tools such as HMMER, SAM and INFERNAL. The most prevalent null models are position-independent residue distributions that include: the uniform distribution, genomic distribution, family-specific distribution and the target sequence distribution. This paper presents a study to evaluate the impact of the choice of a null model in the final result of classifications. In particular, we are interested in minimizing the number of false predictions in a classification. This is a crucial issue to reduce costs of biological validation. Results For all the tests, the target null model presented the lowest number of false positives, when using random sequences as a test. The study was performed in DNA sequences using GC content as the measure of content bias, but the results should be valid also for protein sequences. To broaden the application of the results, the study was performed using randomly generated sequences. Previous studies were performed on aminoacid sequences, using only one probabilistic model (HMM) and on a specific benchmark, and lack more general conclusions about the performance of null models. Finally, a benchmark test with P. falciparum confirmed these results. Conclusions Of the evaluated models the best suited for classification are the uniform model and the target model. However, the use of the uniform model presents a GC bias that can cause more false positives for candidate sequences with extreme compositional bias, a characteristic not described in previous studies. In these cases the target model is more dependable for biological validation due to its higher specificity.

Sequence features responsible for intron retention in human

Abstract Background One of the least common types of alternative splicing is the complete retention of an intron in a mature transcript. Intron retention (IR) is believed to be the result of intron, rather than exon, definition associated with failure of the recognition of weak splice sites flanking short introns. Although studies on individual retained introns have been published, few systematic surveys of large amounts of data have been conducted on the mechanisms that lead to IR. Results TTo understand how sequence features are associated with or control IR, and to produce a generalized model that could reveal previously unknown signals that regulate this type of alternative splicing, we partitioned intron retention events observed in human cDNAs into two groups based on the relative abundance of both isoforms and compared relevant features. We found that a higher frequency of IR in human is associated with individual introns that have weaker splice sites, genes with shorter intron lengths, higher expression levels and lower density of both a set of exon splicing silencers (ESSs) and the intronic splicing enhancer GGG. Both groups of retained introns presented events conserved in mouse, in which the retained introns were also short and presented weaker splice sites. Conclusion Although our results confirmed that weaker splice sites are associated with IR, they showed that this feature alone cannot explain a non-negligible fraction of events. Our analysis suggests that cis-regulatory elements are likely to play a crucial role in regulating IR and also reveals previously unknown features that seem to influence its occurrence. These results highlight the importance of considering the interplay among these features in the regulation of the relative frequency of IR.