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We developed chemically modified PCR primers that allow the design of flexible sticky ends by introducing a photo-cleavable group at the phosphate moiety. Nucleic acid derivatives containing o-nitrobenzyl photo-cleavable groups with a tert-butyl group at the benzyl position were stable during strong base treatment for oligonucleotide synthesis and thermal cycling in PCR reactions. PCR using primers incorporating these nucleic acid derivatives confirmed that chain extension reactions completely stopped at position 1 before and after the site of the photo-cleavable group was introduced. DNA fragments of 2 and 3 kbp, with sticky ends of 50 bases, were successfully concatenated with a high yield of 77%. A plasmid was constructed using this method. Finally, we applied this approach to construct a 48.5 kbp lambda phage DNA, which is difficult to achieve using restriction enzyme-based methods. After 7 days, we were able to confirm the generation of DNA of the desired length. Although the efficiency is yet to be improved, the chemically modified PCR primer offers potential to complement enzymatic methods and serve as a DNA concatenation technique.
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Predicting the secondary structures of RNA molecules is an essential step to characterize their functions, but the thermodynamic probability of any prediction is generally small. On the other hand, there are a few tools for calculating and visualizing various secondary structural information from RNA sequences. We implemented a web server that calculates in parallel various features of secondary structures: different types of secondary structure predictions, the marginal probabilities for local structural contexts, accessibilities of the subsequences, the energy changes by arbitrary base mutations, and the measures for validations of the predicted secondary structures. The web server is available at http://rtools.cbrc.jp , which integrates software tools, CentroidFold, CentroidHomfold, IPknot, CapR, Raccess, Rchange, RintD, and RintW.
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Algoritmos , ARN , Conformación de Ácido Nucleico , ARN/genética , ARN/química , Secuencia de Bases , Análisis de Secuencia de ARN , Programas Informáticos , InternetRESUMEN
Long-read sequencers, such as Pacific Biosciences (PacBio) and Oxford Nanopore Technologies (ONT) sequencers, have improved their read length and accuracy, thereby opening up unprecedented research. Many tools and algorithms have been developed to analyze long reads, and rapid progress in PacBio and ONT has further accelerated their development. Together with the development of high-throughput sequencing technologies and their analysis tools, many read simulators have been developed and effectively utilized. PBSIM is one of the popular long-read simulators. In this study, we developed PBSIM3 with three new functions: error models for long reads, multi-pass sequencing for high-fidelity read simulation and transcriptome sequencing simulation. Therefore, PBSIM3 is now able to meet a wide range of long-read simulation requirements.
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MOTIVATION: Recent advances in high-throughput long-read sequencers, such as PacBio and Oxford Nanopore sequencers, produce longer reads with more errors than short-read sequencers. In addition to the high error rates of reads, non-uniformity of errors leads to difficulties in various downstream analyses using long reads. Many useful simulators, which characterize long-read error patterns and simulate them, have been developed. However, there is still room for improvement in the simulation of the non-uniformity of errors. RESULTS: To capture characteristics of errors in reads for long-read sequencers, here, we introduce a generative model for quality scores, in which a hidden Markov Model with a latest model selection method, called factorized information criteria, is utilized. We evaluated our developed simulator from various points, indicating that our simulator successfully simulates reads that are consistent with real reads. AVAILABILITY AND IMPLEMENTATION: The source codes of PBSIM2 are freely available from https://github.com/yukiteruono/pbsim2. SUPPLEMENTARY INFORMATION: Supplementary data are available at Bioinformatics online.
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Secuenciación de Nucleótidos de Alto Rendimiento , Nanoporos , Simulación por Computador , Análisis de Secuencia de ADN , Programas InformáticosRESUMEN
Methyl-CpG binding protein 2 (MeCP2) is a nuclear protein critical for normal brain function, and both depletion and overexpression of MeCP2 lead to severe neurodevelopmental disease, Rett syndrome (RTT) and MECP2 multiplication disorder, respectively. However, the molecular mechanism by which abnormal MeCP2 dosage causes neuronal dysfunction remains unclear. As MeCP2 expression is nearly equivalent to that of core histones and because it binds DNA throughout the genome, one possible function of MeCP2 is to regulate the 3D structure of chromatin. Here, to examine whether and how MeCP2 levels impact chromatin structure, we used high-resolution confocal and electron microscopy and examined heterochromatic foci of neurons in mice. Using models of RTT and MECP2 triplication syndrome, we found that the heterochromatin structure was significantly affected by the alteration in MeCP2 levels. Analysis of mice expressing either MeCP2-R270X or MeCP2-G273X, which have nonsense mutations in the upstream and downstream regions of the AT-hook 2 domain, respectively, showed that the magnitude of heterochromatin changes was tightly correlated with the phenotypic severity. Postnatal alteration in MeCP2 levels also induced significant changes in the heterochromatin structure, which underscored importance of correct MeCP2 dosage in mature neurons. Finally, functional analysis of MeCP2-overexpressing mice showed that the behavioral and transcriptomic alterations in these mice correlated significantly with the MeCP2 levels and occurred in parallel with the heterochromatin changes. Taken together, our findings demonstrate the essential role of MeCP2 in regulating the 3D structure of neuronal chromatin, which may serve as a potential mechanism that drives pathogenesis of MeCP2-related disorders.SIGNIFICANCE STATEMENT Neuronal function is critically dependent on methyl-CpG binding protein 2 (MeCP2), a nuclear protein abundantly expressed in neurons. The importance of MeCP2 is underscored by the severe childhood neurologic disorders, Rett syndrome (RTT) and MECP2 multiplication disorders, which are caused by depletion and overabundance of MeCP2, respectively. To clarify the molecular function of MeCP2 and to understand the pathogenesis of MECP2-related disorders, we performed detailed structural analyses of neuronal nuclei by using mouse models and high-resolution microscopy. We show that the level of MeCP2 critically regulates 3D structure of heterochromatic foci, and this is mediated in part by the AT-hook 2 domain of MeCP2. Our results demonstrate that one primary function of MeCP2 is to regulate chromatin structure.
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Cromatina/química , Proteína 2 de Unión a Metil-CpG , Neuronas/patología , Estructura Terciaria de Proteína/genética , Animales , Nucléolo Celular/genética , Nucléolo Celular/ultraestructura , Corteza Cerebral/patología , Corteza Cerebral/ultraestructura , Cromatina/ultraestructura , Codón sin Sentido/genética , Discapacidades del Desarrollo/genética , Discapacidades del Desarrollo/patología , Femenino , Histonas/metabolismo , Masculino , Proteína 2 de Unión a Metil-CpG/genética , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Neuronas/ultraestructura , Unión Proteica , Células Piramidales/patología , Células Piramidales/ultraestructura , Transcriptoma/genéticaRESUMEN
Long non-coding RNAs (lncRNAs) play critical roles in various biological processes, but the function of the majority of lncRNAs is still unclear. One approach for estimating a function of a lncRNA is the identification of its interaction target because functions of lncRNAs are expressed through interaction with other biomolecules in quite a few cases. In this paper, we developed "LncRRIsearch," which is a web server for comprehensive prediction of human and mouse lncRNA-lncRNA and lncRNA-mRNA interaction. The prediction was conducted using RIblast, which is a fast and accurate RNA-RNA interaction prediction tool. Users can investigate interaction target RNAs of a particular lncRNA through a web interface. In addition, we integrated tissue-specific expression and subcellular localization data for the lncRNAs with the web server. These data enable users to examine tissue-specific or subcellular localized lncRNA interactions. LncRRIsearch is publicly accessible at http://rtools.cbrc.jp/LncRRIsearch/.
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Summary: LAST-TRAIN improves sequence alignment accuracy by inferring substitution and gap scores that fit the frequencies of substitutions, insertions, and deletions in a given dataset. We have applied it to mapping DNA reads from IonTorrent and PacBio RS, and we show that it reduces reference bias for Oxford Nanopore reads. Availability and Implementation: the source code is freely available at http://last.cbrc.jp/. Contact: mhamada@waseda.jp or mcfrith@edu.k.u-tokyo.ac.jp. Supplementary information: Supplementary data are available at Bioinformatics online.
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Genoma Humano , Polimorfismo Genético , Análisis de Secuencia de ADN/métodos , Programas Informáticos , HumanosRESUMEN
Detection of mutations at the whole-genome level is now possible by the use of high-throughput sequencing. However, determining mutations is a time-consuming process due to the number of false positives provided by mutation-detecting programs. AMAP (automated mutation analysis pipeline) was developed to overcome this issue. AMAP integrates a set of well-validated programs for mapping (BWA), removal of potential PCR duplicates (Picard), realignment (GATK) and detection of mutations (SAMtools, GATK, Pindel, BreakDancer and CNVnator). Thus, all types of mutations such as base substitution, deletion, insertion, translocation and chromosomal rearrangement can be detected by AMAP. In addition, AMAP automatically distinguishes false positives by comparing lists of candidate mutations in sequenced mutants. We tested AMAP by inputting already analyzed read data derived from three individual Arabidopsis thaliana mutants and confirmed that all true mutations were included in the list of candidate mutations. The result showed that the number of false positives was reduced to 12% of that obtained in a previous analysis that lacked a process of reducing false positives. Thus, AMAP will accelerate not only the analysis of mutation induction by individual mutagens but also the process of forward genetics.
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Arabidopsis/genética , Biología Computacional/métodos , Análisis Mutacional de ADN/métodos , Algoritmos , Automatización de Laboratorios , Genoma de Planta , Secuenciación de Nucleótidos de Alto Rendimiento , Polimorfismo de Nucleótido Simple , Análisis de Secuencia de ADN , Programas InformáticosRESUMEN
The secondary structures, as well as the nucleotide sequences, are the important features of RNA molecules to characterize their functions. According to the thermodynamic model, however, the probability of any secondary structure is very small. As a consequence, any tool to predict the secondary structures of RNAs has limited accuracy. On the other hand, there are a few tools to compensate the imperfect predictions by calculating and visualizing the secondary structural information from RNA sequences. It is desirable to obtain the rich information from those tools through a friendly interface. We implemented a web server of the tools to predict secondary structures and to calculate various structural features based on the energy models of secondary structures. By just giving an RNA sequence to the web server, the user can get the different types of solutions of the secondary structures, the marginal probabilities such as base-paring probabilities, loop probabilities and accessibilities of the local bases, the energy changes by arbitrary base mutations as well as the measures for validations of the predicted secondary structures. The web server is available at http://rtools.cbrc.jp, which integrates software tools, CentroidFold, CentroidHomfold, IPKnot, CapR, Raccess, Rchange and RintD.
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Conformación de Ácido Nucleico , Pliegue del ARN , ARN/química , Programas Informáticos , Algoritmos , Emparejamiento Base , Secuencia de Bases , Gráficos por Computador , Internet , Mutación , ARN/genética , Análisis de Secuencia de ARN , TermodinámicaRESUMEN
MOTIVATION: Recent studies have suggested that both the genome and the genome with epigenetic modifications, the so-called epigenome, play important roles in various biological functions, such as transcription and DNA replication, repair, and recombination. It is well known that specific combinations of histone modifications (e.g. methylations and acetylations) of nucleosomes induce chromatin states that correspond to specific functions of chromatin. Although the advent of next-generation sequencing (NGS) technologies enables measurement of epigenetic information for entire genomes at high-resolution, the variety of chromatin states has not been completely characterized. RESULTS: In this study, we propose a method to estimate the chromatin states indicated by genome-wide chromatin marks identified by NGS technologies. The proposed method automatically estimates the number of chromatin states and characterize each state on the basis of a hidden Markov model (HMM) in combination with a recently proposed model selection technique, factorized information criteria. The method is expected to provide an unbiased model because it relies on only two adjustable parameters and avoids heuristic procedures as much as possible. Computational experiments with simulated datasets show that our method automatically learns an appropriate model, even in cases where methods that rely on Bayesian information criteria fail to learn the model structures. In addition, we comprehensively compare our method to ChromHMM on three real datasets and show that our method estimates more chromatin states than ChromHMM for those datasets.
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Cromatina/metabolismo , Epigenómica/métodos , Epigénesis Genética , Secuenciación de Nucleótidos de Alto Rendimiento , Humanos , Cadenas de Markov , Nucleosomas/metabolismoRESUMEN
MOTIVATION: PacBio sequencers produce two types of characteristic reads (continuous long reads: long and high error rate and circular consensus sequencing: short and low error rate), both of which could be useful for de novo assembly of genomes. Currently, there is no available simulator that targets the specific generation of PacBio libraries. RESULTS: Our analysis of 13 PacBio datasets showed characteristic features of PacBio reads (e.g. the read length of PacBio reads follows a log-normal distribution). We have developed a read simulator, PBSIM, that captures these features using either a model-based or sampling-based method. Using PBSIM, we conducted several hybrid error correction and assembly tests for PacBio reads, suggesting that a continuous long reads coverage depth of at least 15 in combination with a circular consensus sequencing coverage depth of at least 30 achieved extensive assembly results. AVAILABILITY: PBSIM is freely available from the web under the GNU GPL v2 license (http://code.google.com/p/pbsim/).
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Genómica/métodos , Análisis de Secuencia de ADN/métodos , Programas InformáticosRESUMEN
PIWI-interacting RNAs (piRNAs) silence retrotransposons in Drosophila germ lines by associating with the PIWI proteins Argonaute 3 (AGO3), Aubergine (Aub) and Piwi. piRNAs in Drosophila are produced from intergenic repetitive genes and piRNA clusters by two systems: the primary processing pathway and the amplification loop. The amplification loop occurs in a Dicer-independent, PIWI-Slicer-dependent manner. However, primary piRNA processing remains elusive. Here we analysed piRNA processing in a Drosophila ovarian somatic cell line where Piwi, but not Aub or AGO3, is expressed; thus, only the primary piRNAs exist. In addition to flamenco, a Piwi-specific piRNA cluster, traffic jam (tj), a large Maf gene, was determined as a new piRNA cluster. piRNAs arising from tj correspond to the untranslated regions of tj messenger RNA and are sense-oriented. piRNA loading on to Piwi may occur in the cytoplasm. zucchini, a gene encoding a putative cytoplasmic nuclease, is required for tj-derived piRNA production. In tj and piwi mutant ovaries, somatic cells fail to intermingle with germ cells and Fasciclin III is overexpressed. Loss of tj abolishes Piwi expression in gonadal somatic cells. Thus, in gonadal somatic cells, tj gives rise simultaneously to two different molecules: the TJ protein, which activates Piwi expression, and piRNAs, which define the Piwi targets for silencing.
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Proteínas de Drosophila/metabolismo , Drosophila melanogaster/metabolismo , Factores de Transcripción Maf de Gran Tamaño/metabolismo , Proteínas Proto-Oncogénicas/metabolismo , Complejo Silenciador Inducido por ARN/metabolismo , ARN/metabolismo , Animales , Proteínas Argonautas , Moléculas de Adhesión Celular Neuronal/metabolismo , Línea Celular , Proteínas de Drosophila/genética , Drosophila melanogaster/genética , Endorribonucleasas/metabolismo , Femenino , Genes de Insecto/genética , Sitios Genéticos/genética , Factores de Transcripción Maf de Gran Tamaño/genética , Masculino , Ovario/citología , Ovario/metabolismo , Fenotipo , Proteínas Proto-Oncogénicas/genética , ARN/biosíntesis , ARN/genética , Interferencia de ARN , Procesamiento Postranscripcional del ARN , Complejo Silenciador Inducido por ARN/genética , Testículo/citología , Testículo/metabolismoRESUMEN
In order to support high-throughput screening for ligands of G-protein coupled receptors (GPCRs) by using bioinformatics technology, we introduce a database (SEVENS) with genome-scale annotation and software (GRIFFIN) that can simulate GPCR function. SEVENS ( http://sevens.cbrc.jp/ ) is an integrated database that includes GPCR genes that are identified with high accuracy (99.4% sensitivity and 96.6% specificity) from various types of genomes, by a pipeline that integrates such software as a gene finder, a sequence alignment tool, a motif and domain assignment tool, and a transmembrane helix (TMH) predictor. SEVENS provides the user a genome-scale overview of the "GPCR universe" with detailed information of chromosomal mapping, phylogenetic tree, protein sequence and structure, and experimental evidence, all of which are accessible via a user-friendly interface. GRIFFIN ( http://griffin.cbrc.jp/ ) can predict GPCR and G-protein coupling selectivity induced by ligand binding with high sensitivity and specificity (more than 87% on average), based on the support vector machine (SVM) and hidden Markov Model (HMM). SEVENS and GRIFFIN are expected to contribute to revealing the function of orphan and unknown GPCRs.
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Bases de Datos Genéticas , Genómica/métodos , Receptores Acoplados a Proteínas G/genética , Animales , Inteligencia Artificial , Biología Computacional , Diseño de Fármacos , Evaluación Preclínica de Medicamentos/métodos , Evaluación Preclínica de Medicamentos/estadística & datos numéricos , Genómica/estadística & datos numéricos , Ensayos Analíticos de Alto Rendimiento/estadística & datos numéricos , Humanos , Ligandos , Cadenas de Markov , Proteómica/métodos , Proteómica/estadística & datos numéricos , Receptores Acoplados a Proteínas G/química , Receptores Acoplados a Proteínas G/metabolismo , Programas InformáticosRESUMEN
BACKGROUND: The olfactory system plays an important role in the recognition of leaf volatiles during the search of folivore insects for a suitable plant host. For example, volatiles emitted by mulberry leaves trigger chemotaxis behavior in the silkworms Bombyx mori, and as a consequence, they preferentially reside on and consume mulberry leaves. Here, we aimed to identify natural chemoattractants and their corresponding olfactory receptors (Ors) involved in silkworm behavior to mulberry leaves. RESULTS: Chemotaxis behavioral assays for headspace volatiles detected by gas chromatography-mass spectroscopy analysis revealed that among the volatiles that were emitted by mulberry leaves, cis-jasmone was the most potent attractant for silkworms, working at a threshold of 30 pg from [corrected] 20 cm distance. Among a total of 66 Ors identified in the B. mori genome, we found that 23 were expressed in the olfactory organs during larval stages. Functional analysis of all the larvae-expressed Ors in Xenopus oocytes revealed that one Or, termed BmOr-56, showed a high sensitivity to cis-jasmone. In addition, the ligand-receptor activity of BmOr-56 reflected the chemotaxis behavioral response of silkworms. CONCLUSIONS: We identified cis-jasmone as a potent attractant in mulberry leaves for silkworms and provide evidence that a highly tuned receptor, BmOr-56, may mediate this behavioral attraction. The current study sheds light on the mechanism of the correlation between olfactory perception in folivore insects and chemotaxis behavior to a natural volatile emitted by green leaves.
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Bombyx/fisiología , Factores Quimiotácticos/química , Morus/química , Aceites Volátiles/química , Receptores Odorantes/fisiología , Animales , Bombyx/efectos de los fármacos , Bombyx/genética , Fraccionamiento Químico , Factores Quimiotácticos/aislamiento & purificación , Quimiotaxis/efectos de los fármacos , Cromatografía de Gases , Conducta Alimentaria , Larva/efectos de los fármacos , Larva/fisiología , Filogenia , Hojas de la Planta/química , Receptores Odorantes/genética , Relación Estructura-Actividad , XenopusRESUMEN
We developed a pair of databases that support two important tasks: annotation of anonymous RNA transcripts and discovery of novel non-coding RNAs. The database combo is called the Functional RNA Database and consists of two databases: a rewrite of the original version of the Functional RNA Database (fRNAdb) and the latest version of the UCSC GenomeBrowser for Functional RNA. The former is a sequence database equipped with a powerful search function and hosts a large collection of known/predicted non-coding RNA sequences acquired from existing databases as well as novel/predicted sequences reported by researchers of the Functional RNA Project. The latter is a UCSC Genome Browser mirror with large additional custom tracks specifically associated with non-coding elements. It also includes several functional enhancements such as a presentation of a common secondary structure prediction at any given genomic window < or =500 bp. Our GenomeBrowser supports user authentication and user-specific tracks. The current version of the fRNAdb is a complete rewrite of the former version, hosting a larger number of sequences and with a much friendlier interface. The current version of UCSC GenomeBrowser for Functional RNA features a larger number of tracks and richer features than the former version. The databases are available at http://www.ncrna.org/.
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Bases de Datos de Ácidos Nucleicos , ARN no Traducido/química , Animales , Genómica , Humanos , Ratones , ARN no Traducido/fisiología , Ratas , Análisis de Secuencia de ARNRESUMEN
RNA silencing is a conserved mechanism in which small RNAs trigger various forms of sequence-specific gene silencing by guiding Argonaute complexes to target RNAs by means of base pairing. RNA silencing is thought to have evolved as a form of nucleic-acid-based immunity to inactivate viruses and transposable elements. Although the activity of transposable elements in animals has been thought largely to be restricted to the germ line, recent studies have shown that they may also actively transpose in somatic cells, creating somatic mosaicism in animals. In the Drosophila germ line, Piwi-interacting RNAs arise from repetitive intergenic elements including retrotransposons by a Dicer-independent pathway and function through the Piwi subfamily of Argonautes to ensure silencing of retrotransposons. Here we show that, in cultured Drosophila S2 cells, Argonaute 2 (AGO2), an AGO subfamily member of Argonautes, associates with endogenous small RNAs of 20-22 nucleotides in length, which we have collectively named endogenous short interfering RNAs (esiRNAs). esiRNAs can be divided into two groups: one that mainly corresponds to a subset of retrotransposons, and the other that arises from stem-loop structures. esiRNAs are produced in a Dicer-2-dependent manner from distinctive genomic loci, are modified at their 3' ends and can direct AGO2 to cleave target RNAs. Mutations in Dicer-2 caused an increase in retrotransposon transcripts. Together, our findings indicate that different types of small RNAs and Argonautes are used to repress retrotransposons in germline and somatic cells in Drosophila.
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Proteínas de Drosophila/metabolismo , Drosophila melanogaster/citología , Drosophila melanogaster/metabolismo , ARN Interferente Pequeño/metabolismo , Complejo Silenciador Inducido por ARN/metabolismo , Animales , Proteínas Argonautas , Línea Celular , Proteínas de Drosophila/genética , Drosophila melanogaster/enzimología , Drosophila melanogaster/genética , Factores Eucarióticos de Iniciación , Células Germinativas/metabolismo , Mosaicismo , Reacción en Cadena de la Polimerasa , Unión Proteica , ARN Helicasas/genética , ARN Helicasas/metabolismo , Interferencia de ARN , ARN Interferente Pequeño/genética , Retroelementos/genética , Ribonucleasa IIIRESUMEN
SUMMARY: We have launched a web server, which serves as a general-purpose idiogram rendering service, and allows users to generate high-quality idiograms with custom annotation according to their own genome-wide mapping/annotation data through an easy-to-use interface. The generated idiograms are suitable not only for visualizing summaries of genome-wide analysis but also for many types of presentation material including web pages, conference posters, oral presentations, etc. AVAILABILITY: Idiographica is freely available at http://www.ncrna.org/idiographica/
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Algoritmos , Bandeo Cromosómico/métodos , Mapeo Cromosómico/métodos , Gráficos por Computador , Internet , Programas Informáticos , Interfaz Usuario-Computador , Animales , Humanos , Ratones , RatasRESUMEN
There are abundance of transcripts that code for no particular protein and that remain functionally uncharacterized. Some of these transcripts may have novel functions while others might be junk transcripts. Unfortunately, the experimental validation of such transcripts to find functional non-coding RNA candidates is very costly. Therefore, our primary interest is to computationally mine candidate functional transcripts from a pool of uncharacterized transcripts. We introduce fRNAdb: a novel database service that hosts a large collection of non-coding transcripts including annotated/non-annotated sequences from the H-inv database, NONCODE and RNAdb. A set of computational analyses have been performed on the included sequences. These analyses include RNA secondary structure motif discovery, EST support evaluation, cis-regulatory element search, protein homology search, etc. fRNAdb provides an efficient interface to help users filter out particular transcripts under their own criteria to sort out functional RNA candidates. fRNAdb is available at http://www.ncrna.org/
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Bases de Datos de Ácidos Nucleicos , ARN no Traducido/química , Secuencia de Bases , Genómica , Internet , MicroARNs/fisiología , ARN Mensajero/química , ARN no Traducido/fisiología , Interfaz Usuario-ComputadorRESUMEN
We have developed an automatic system for identifying GPCR (G-protein coupled receptor) genes from various kinds of genomes, which is finally deposited in the SEVENS database (http://sevens.cbrc.jp/), by integrating such software as a gene finder, a sequence alignment tool, a motif and domain assignment tool, and a transmembrane helix predictor. SEVENS enables us to perform a genome-scale overview of the "GPCR universe" using sequences that are identified with high accuracy (99.4% sensitivity and 96.6% specificity). Using this system, we surveyed the complete genomes of 7 eukaryotes and 224 prokaryotes, and found that there are 4 to 1016 GPCR genes in the 7 eukaryotes, and only a total of 16 GPCR genes in all the prokaryotes. Our preliminary results indicate that 11 subfamilies of the Class A family, the Class 2(B) family, the Class 3(C) family and the fz/smo family are commonly found among human, fly, and nematode genomes. We also analyzed the chromosomal locations of the GPCR genes with the Kolmogorov-Smirnov test, and found that species-specific families, such as olfactory, taste, and chemokine receptors in human and nematode chemoreceptor in worm, tend to form clusters extensively, whereas no significant clusters were detected in fly and plant genomes.