MPBLAST : improved BLAST performance with multiplexed queries

SUMMARY: We have developed a program, MPBLAST, that increases the throughput of batch BLASTN searches by multiplexing (concatenating) query sequences and thereby reducing the number of actual database searches performed. Throughput was observed to increase in reciprocal proportion to the component sequence length. For sequencing read-sized queries of 500 bp, an order of magnitude speed-up was seen. AVAILABILITY: Free (see http://blast.wustl.edu) CONTACT: [ikorf, gish]@watson.wustl.edu     

Pfam 3.1: 1313 multiple alignments and profile HMMs match the majority of proteins.

Pfam is a collection of multiple alignments and profile hidden Markov models of protein domain families. Release 3.1 is a major update of the Pfam database and contains 1313 families which are available on the World Wide Web in Europe at http://www.sanger.ac.uk/Software/Pfam/ and http://www.cgr.ki.se/Pfam/, and in the US at http://pfam.wustl.edu/. Over 54% of proteins in SWISS-PROT-35 and SP-TrEMBL-5 match a Pfam family. The primary changes of Pfam since release 2.1 are that we now use the more advanced version 2 of the HMMER software, which is more sensitive and provides expectation values for matches, and that it now includes proteins from both SP-TrEMBL and SWISS-PROT.     

WU-Blast2 server at the European Bioinformatics Institute

Since 1995, the WU-BLAST programs (http://blast.wustl.edu) have provided a fast, flexible and reliable method for similarity searching of biological sequence databases. The software is in use at many locales and web sites. The European Bioinformatics Institute's WU-Blast2 (http://www.ebi.ac.uk/blast2/) server has been providing free access to these search services since 1997 and today supports many features that both enhance the usability and expand on the scope of the software.     

Procom: a web-based tool to compare multiple eukaryotic proteomes

SUMMARY: Each organism has traits that are shared with some, but not all, organisms. Identification of genes needed for a particular trait can be accomplished by a comparative genomics approach using three or more organisms. Genes that occur in organisms without the trait are removed from the set of genes in common among organisms with the trait. To facilitate these comparisons, a web-based server, Procom, was developed to identify the subset of genes that may be needed for a trait. AVAILABILITY: The Procom program is freely available with documentation and examples at http://ural.wustl.edu/~billy/Procom/ CONTACT: billy@ural.wustl.edu.     

An iterated loop matching approach to the prediction of RNA secondary structures with pseudoknots

MOTIVATION: Pseudoknots have generally been excluded from the prediction of RNA secondary structures due to its difficulty in modeling. Although, several dynamic programming algorithms exist for the prediction of pseudoknots using thermodynamic approaches, they are neither reliable nor efficient. On the other hand, comparative methods are more reliable, but are often done in an ad hoc manner and require expert intervention. Maximum weighted matching, an algorithm for pseudoknot prediction with comparative analysis, suffers from low-prediction accuracy in many cases. RESULTS: Here we present an algorithm, iterated loop matching, for reliably and efficiently predicting RNA secondary structures including pseudoknots. The method can utilize either thermodynamic or comparative information or both, thus is able to predict pseudoknots for both aligned and individual sequences. We have tested the algorithm on a number of RNA families. Using 8-12 homologous sequences, the algorithm correctly identifies more than 90% of base-pairs for short sequences and 80% overall. It correctly predicts nearly all pseudoknots and produces very few spurious base-pairs for sequences without pseudoknots. Comparisons show that our algorithm is both more sensitive and more specific than the maximum weighted matching method. In addition, our algorithm has high-prediction accuracy on individual sequences, comparable with the PKNOTS algorithm, while using much less computational resources. AVAILABILITY: The program has been implemented in ANSI C and is freely available for academic use at http://www.cse.wustl.edu/~zhang/projects/rna/ilm/ Supplementary information: http://www.cse.wustl.edu/~zhang/projects/rna/ilm/     

SPrCY: comparison of structural predictions in the Saccharomyces cerevisiae genome

SUMMARY: SPrCY is a web-accessible database which provides comparison of structure prediction results for the Saccharomyces cerevisiae genome. This web service offers the ability to search, analyze and compare the yeast structural predictions from sequence-only (Superfamily, PDBAA BLAST and Pfam) and sequence-structure-based (SAM-T02, 3D-PSSM, mGenTHREADER) methods. AVAILABILITY: The service is freely available via web at http://agave.wustl.edu/yeast/     

Rfam: an RNA family database

Rfam is a collection of multiple sequence alignments and covariance models representing non-coding RNA families. Rfam is available on the web in the UK at http://www.sanger.ac.uk/Software/Rfam/ and in the US at http://rfam.wustl.edu/. These websites allow the user to search a query sequence against a library of covariance models, and view multiple sequence alignments and family annotation. The database can also be downloaded in flatfile form and searched locally using the INFERNAL package (http://infernal.wustl.edu/). The first release of Rfam (1.0) contains 25 families, which annotate over 50 000 non-coding RNA genes in the taxonomic divisions of the EMBL nucleotide database.     

The Pfam protein families database

Pfam is a large collection of protein multiple sequence alignments and profile hidden Markov models. Pfam is available on the WWW in the UK at http://www.sanger.ac.uk/Software/Pfam/, in Sweden at http://www.cgr.ki.se/Pfam/ and in the US at http://pfam.wustl.edu/. The latest version (4.3) of Pfam contains 1815 families. These Pfam families match 63% of proteins in SWISS-PROT 37 and TrEMBL 9. For complete genomes Pfam currently matches up to half of the proteins. Genomic DNA can be directly searched against the Pfam library using the Wise2 package.     

Using mRNAs lengths to accurately predict the alternatively spliced gene products in Caenorhabditis elegans

MOTIVATION: Computational gene prediction methods are an important component of whole genome analyses. While ab initio gene finders have demonstrated major improvements in accuracy, the most reliable methods are evidence-based gene predictors. These algorithms can rely on several different sources of evidence including predictions from multiple ab initio gene finders, matches to known proteins, sequence conservation and partial cDNAs to predict the final product. Despite the success of these algorithms, prediction of complete gene structures, especially for alternatively spliced products, remains a difficult task. RESULTS: LOCUS (Length Optimized Characterization of Unknown Spliceforms) is a new evidence-based gene finding algorithm which integrates a length-constraint into a dynamic programming-based framework for prediction of gene products. On a Caenorhabditis elegans test set of alternatively spliced internal exons, its performance exceeds that of current ab initio gene finders and in most cases can accurately predict the correct form of all the alternative products. As the length information used by the algorithm can be obtained in a high-throughput fashion, we propose that integration of such information into a gene-prediction pipeline is feasible and doing so may improve our ability to fully characterize the complete set of mRNAs for a genome. AVAILABILITY: LOCUS is available from http://ural.wustl.edu/software.html     

Pfam: multiple sequence alignments and HMM-profiles of protein domains.

Pfam contains multiple alignments and hidden Markov model based profiles (HMM-profiles) of complete protein domains. The definition of domain boundaries, family members and alignment is done semi-automatically based on expert knowledge, sequence similarity, other protein family databases and the ability of HMM-profiles to correctly identify and align the members. Release 2.0 of Pfam contains 527 manually verified families which are available for browsing and on-line searching via the World Wide Web in the UK at http://www.sanger.ac.uk/Pfam/ and in the US at http://genome.wustl. edu/Pfam/ Pfam 2.0 matches one or more domains in 50% of Swissprot-34 sequences, and 25% of a large sample of predicted proteins from the Caenorhabditis elegans genome.     

The Treeterbi and Parallel Treeterbi algorithms: efficient, optimal decoding for ordinary, generalized and pair HMMs

MOTIVATION: Hidden Markov models (HMMs) and generalized HMMs been successfully applied to many problems, but the standard Viterbi algorithm for computing the most probable interpretation of an input sequence (known as decoding) requires memory proportional to the length of the sequence, which can be prohibitive. Existing approaches to reducing memory usage either sacrifice optimality or trade increased running time for reduced memory. RESULTS: We developed two novel decoding algorithms, Treeterbi and Parallel Treeterbi, and implemented them in the TWINSCAN/N-SCAN gene-prediction system. The worst case asymptotic space and time are the same as for standard Viterbi, but in practice, Treeterbi optimally decodes arbitrarily long sequences with generalized HMMs in bounded memory without increasing running time. Parallel Treeterbi uses the same ideas to split optimal decoding across processors, dividing latency to completion by approximately the number of available processors with constant average overhead per processor. Using these algorithms, we were able to optimally decode all human chromosomes with N-SCAN, which increased its accuracy relative to heuristic solutions. We also implemented Treeterbi for Pairagon, our pair HMM based cDNA-to-genome aligner. AVAILABILITY: The TWINSCAN/N-SCAN/PAIRAGON open source software package is available from http://genes.cse.wustl.edu.     

RNA Sampler: a new sampling based algorithm for common RNA secondary structure prediction and structural alignment

MOTIVATION: Non-coding RNA genes and RNA structural regulatory motifs play important roles in gene regulation and other cellular functions. They are often characterized by specific secondary structures that are critical to their functions and are often conserved in phylogenetically or functionally related sequences. Predicting common RNA secondary structures in multiple unaligned sequences remains a challenge in bioinformatics research. Methods and RESULTS: We present a new sampling based algorithm to predict common RNA secondary structures in multiple unaligned sequences. Our algorithm finds the common structure between two sequences by probabilistically sampling aligned stems based on stem conservation calculated from intrasequence base pairing probabilities and intersequence base alignment probabilities. It iteratively updates these probabilities based on sampled structures and subsequently recalculates stem conservation using the updated probabilities. The iterative process terminates upon convergence of the sampled structures. We extend the algorithm to multiple sequences by a consistency-based method, which iteratively incorporates and reinforces consistent structure information from pairwise comparisons into consensus structures. The algorithm has no limitation on predicting pseudoknots. In extensive testing on real sequence data, our algorithm outperformed other leading RNA structure prediction methods in both sensitivity and specificity with a reasonably fast speed. It also generated better structural alignments than other programs in sequences of a wide range of identities, which more accurately represent the RNA secondary structure conservations. AVAILABILITY: The algorithm is implemented in a C program, RNA Sampler, which is available at http://ural.wustl.edu/software.html     

The Pfam Protein Families Database

Pfam is a large collection of protein multiple sequence alignments and profile hidden Markov models. Pfam is available on the World Wide Web in the UK at http://www.sanger.ac.uk/Software/Pfam/, in Sweden at http://www.cgb.ki.se/Pfam/, in France at http://pfam.jouy.inra.fr/ and in the US at http://pfam.wustl.edu/. The latest version (6.6) of Pfam contains 3071 families, which match 69% of proteins in SWISS-PROT 39 and TrEMBL 14. Structural data, where available, have been utilised to ensure that Pfam families correspond with structural domains, and to improve domain-based annotation. Predictions of non-domain regions are now also included. In addition to secondary structure, Pfam multiple sequence alignments now contain active site residue mark-up. New search tools, including taxonomy search and domain query, greatly add to the functionality and usability of the Pfam resource.     

PROBER: oligonucleotide FISH probe design software

PROBER is an oligonucleotide primer design software application that designs multiple primer pairs for generating PCR probes useful for fluorescence in situ hybridization (FISH). PROBER generates Tiling Oligonucleotide Probes (TOPs) by masking repetitive genomic sequences and delineating essentially unique regions that can be amplified to yield small (100-2000 bp) DNA probes that in aggregate will generate a single, strong fluorescent signal for regions as small as a single gene. TOPs are an alternative to bacterial artificial chromosomes (BACs) that are commonly used for FISH but may be unstable, unavailable, chimeric, or non-specific to small (10-100 kb) genomic regions. PROBER can be applied to any genomic locus, with the limitation that the locus must contain at least 10 kb of essentially unique blocks. To test the software, we designed a number of probes for genomic amplifications and hemizygous deletions that were initially detected by Representational Oligonucleotide Microarray Analysis of breast cancer tumors. AVAILABILITY: http://prober.cshl.edu     

ToxoDB: accessing the Toxoplasma gondii genome

ToxoDB (http://ToxoDB.org) provides a genome resource for the protozoan parasite Toxoplasma gondii. Several sequencing projects devoted to T. gondii have been completed or are in progress: an EST project (http://genome.wustl.edu/est/index.php?toxoplasma=1), a BAC clone end-sequencing project (http://www.sanger.ac.uk/Projects/T_gondii/) and an 8X random shotgun genomic sequencing project (http://www.tigr.org/tdb/e2k1/tga1/). ToxoDB was designed to provide a central point of access for all available T. gondii data, and a variety of data mining tools useful for the analysis of unfinished, un-annotated draft sequence during the early phases of the genome project. In later stages, as more and different types of data become available (microarray, proteomic, SNP, QTL, etc.) the database will provide an integrated data analysis platform facilitating user-defined queries across the different data types.     

Applications of Tree-Maps to hierarchical biological data

SUMMARY: A brief overview of Tree-Maps provides the basis for understanding two new implementations of Tree-Map methods. TreeMapClusterView provides a new way to view microarray gene expression data, and GenePlacer provides a view of gene ontology annotation data. We also discuss the benefits of Tree-Maps to visualize complex hierarchies in functional genomics. AVAILABILITY: Java class files are freely available at http://mendel.mc.duke.edu/bioinformatics/ CONTACT: mccon012@mc.duke.edu SUPPLEMENTARY INFORMATION: For more information on TreeMapClusterView (see http://mendel.mc.duke.edu/bioinformatics/software/boxclusterview/), and http://mendel.mc.duke.edu/bioinformatics/software/geneplacer/).     

A simple algorithm to infer gene duplication and speciation events on a gene tree

MOTIVATION: When analyzing protein sequences using sequence similarity searches, orthologous sequences (that diverged by speciation) are more reliable predictors of a new protein's function than paralogous sequences (that diverged by gene duplication), because duplication enables functional diversification. The utility of phylogenetic information in high-throughput genome annotation ('phylogenomics') is widely recognized, but existing approaches are either manual or indirect (e.g. not based on phylogenetic trees). Our goal is to automate phylogenomics using explicit phylogenetic inference. A necessary component is an algorithm to infer speciation and duplication events in a given gene tree. RESULTS: We give an algorithm to infer speciation and duplication events on a gene tree by comparison to a trusted species tree. This algorithm has a worst-case running time of O(n(2)) which is inferior to two previous algorithms that are approximately O(n) for a gene tree of sequences. However, our algorithm is extremely simple, and its asymptotic worst case behavior is only realized on pathological data sets. We show empirically, using 1750 gene trees constructed from the Pfam protein family database, that it appears to be a practical (and often superior) algorithm for analyzing real gene trees. AVAILABILITY: http://www.genetics.wustl.edu/eddy/forester.     

Thermodynamic parameters for DNA sequences with dangling ends

The thermodynamic contributions to duplex formation of all 32 possible single-nucleotide dangling ends on a Watson-Crick pair are reported. In most instances, dangling ends are stabilizing with free energy contributions ranging from +0.48 (GT(A)) to-0.96 kcal/mol (). In comparison, Watson-Crick nearest-neighbor increments range from -0. 58 (TA/AT) to -2.24 (GC/CG) kcal/mol. Hence, in some cases, a dangling end contributes as much to duplex stability as a Watson-Crick A-T base pair. The implications of these results for DNA probe design are discussed. Analysis of the sequence dependence of dangling-end stabilities show that the nature of the closing base pair largely determines the stabilization. For a given closing base pair, however, adenine dangling ends are always more or equally as stable as the other dangling nucleotides. Moreover, 5' dangling ends are more or equally as stabilizing as their 3' counterparts. Comparison of DNA with RNA dangling-end motifs shows that DNA motifs with 5' dangling ends contribute to stability equally or more than their RNA counterparts. Conversely, RNA 3' dangling ends contribute to stability equally or more than their DNA counterparts. This data set has been incorporated into a DNA secondary structure prediction algorithm (DNA MFOLD) (http://mfold2.wustl.edu/mfold/dna/for m1.cgi) as well as a DNA hybridization prediction algorithm (HYTHERtrade mark) (http://jsl1.chem.wayne.edu/Hyther/hythermenu .html).