REBASE--restriction enzymes and methylases.

REBASE is a comprehensive database of information about restriction enzymes and their associated methylases, including their recognition and cleavage sites and their commercial availability. Information from REBASE is available via monthly electronic mailings as well as via WAIS, anonymous ftp and through the World Wide Web (http://www.neb.com/rebase). Specialized files are available that can be used directly by many software packages.     

REBASE--restriction enzymes and methylases.

REBASE is a comprehensive database of information about restriction enzymes and their associated methylases, including their recognition and cleavage sites and their commercial availability. Information from REBASE is available via monthly electronic mailings as well as via WAIS and anonymous ftp. Specialized files are available that can be used directly by many software packages.     

NEBcutter: A program to cleave DNA with restriction enzymes

NEBcutter, version 1.0, is a program available via a web server (http://tools.neb.com/NEBcutter) that will accept an input DNA sequence and produce a comprehensive report of the restriction enzymes that will cleave the sequence. It produces a variety of outputs including restriction enzyme maps, theoretical digests and links into the restriction enzyme database, REBASE (http://www.neb.com/rebase). Importantly, its table of recognition sites is updated daily from REBASE and it marks all sites that are potentially affected by DNA methylation (Dam, Dcm, etc.). Many options exist to choose the enzymes used for digestion, including all known specificities, subsets of those that are commercially available or sets of enzymes that produce compatible termini.     

REBASE: restriction enzymes and methyltransferases

REBASE contains comprehensive information about restriction enzymes, DNA methyltransferases and related proteins such as nicking enzymes, specificity subunits and control proteins. It contains published and unpublished references, recognition and cleavage sites, isoschizomers, commercial availability, crystal and sequence data. Homing endonucleases are also included. REBASE contains the most complete and up-to-date information about the methylation sensitivity of restriction endonucleases. In addition, there is extensive information about the known and putative restriction-modification (R-M) systems in more than 100 sequenced bacterial and archaeal genomes. The data is available on the web (http://rebase.neb.com/rebase/rebase.html), through ftp (ftp.neb.com) and as monthly updates via email.     

Structural and evolutionary classification of Type II restriction enzymes based on theoretical and experimental analyses

For a very long time, Type II restriction enzymes (REases) have been a paradigm of ORFans: proteins with no detectable similarity to each other and to any other protein in the database, despite common cellular and biochemical function. Crystallographic analyses published until January 2008 provided high-resolution structures for only 28 of 1637 Type II REase sequences available in the Restriction Enzyme database (REBASE). Among these structures, all but two possess catalytic domains with the common PD-(D/E)XK nuclease fold. Two structures are unrelated to the others: R.BfiI exhibits the phospholipase D (PLD) fold, while R.PabI has a new fold termed 'half-pipe'. Thus far, bioinformatic studies supported by site-directed mutagenesis have extended the number of tentatively assigned REase folds to five (now including also GIY-YIG and HNH folds identified earlier in homing endonucleases) and provided structural predictions for dozens of REase sequences without experimentally solved structures. Here, we present a comprehensive study of all Type II REase sequences available in REBASE together with their homologs detectable in the nonredundant and environmental samples databases at the NCBI. We present the summary and critical evaluation of structural assignments and predictions reported earlier, new classification of all REase sequences into families, domain architecture analysis and new predictions of three-dimensional folds. Among 289 experimentally characterized (not putative) Type II REases, whose apparently full-length sequences are available in REBASE, we assign 199 (69%) to contain the PD-(D/E)XK domain. The HNH domain is the second most common, with 24 (8%) members. When putative REases are taken into account, the fraction of PD-(D/E)XK and HNH folds changes to 48% and 30%, respectively. Fifty-six characterized (and 521 predicted) REases remain unassigned to any of the five REase folds identified so far, and may exhibit new architectures. These enzymes are proposed as the most interesting targets for structure determination by high-resolution experimental methods. Our analysis provides the first comprehensive map of sequence-structure relationships among Type II REases and will help to focus the efforts of structural and functional genomics of this large and biotechnologically important class of enzymes.     

REBASE - restriction enzymes and methylases

REBASE is a comprehensive database of information about restriction enzymes and related proteins. It contains published and unpublished references, recognition and cleavage sites, isoschizomers, commercial availability, methylation sensitivity, crystal and sequence data. DNA methyltransferases, homing endonucleases, nicking enzymes, specificity subunits and control proteins are also included. Most recently, putative DNA methyltransferases and restriction enzymes, as predicted from analysis of genomic sequences, are also listed. The data is distributed via Email, ftp (ftp.neb.com), and the Web (http://rebase.neb.com).     

The biology of restriction and anti-restriction

The phenomena of prokaryotic restriction and modification, as well as anti-restriction, were first discovered five decades ago but have yielded only gradually to rigorous analysis. Work presented at the 5th New England Biolabs Meeting on Restriction-Modification (available on REBASE, http://www.rebase.com) and several recently published genetic, biochemical and biophysical analyses indicate that these fields continue to contribute significantly to basic science. Recently, there have been several studies that have shed light on the still developing field of restriction-modification and on the newly re-emerging field of anti-restriction.     

REBASE--restriction enzymes and methylases

REBASE contains comprehensive information about restriction enzymes, DNA methylases and related proteins such as nicking enzymes, specificity subunits and control proteins. It contains published and unpublished references, recognition and cleavage sites, isoschizomers, commercial availability, methylation sensitivity, crystal data and sequence data. Homing endonucleases are also included. Most recently, extensive information about the methylation sensitivity of restriction enzymes has been added and a new feature contains complete analyses of the putative restriction systems in the sequenced bacterial and archaeal genomes. The data is distributed via email, ftp (ftp.neb.com) and the Web (http://rebase. neb.com).     

Analysis of bacterial RM-systems through genome-scale analysis and related taxonomy issues

Recognition sites for type II restriction and modification enzymes in genomes of several bacteria are recognized as semi-palindromic motifs and are avoided at a significant degree. The key idea of contrast word analysis with respect to RMS recognition sites, is that under-represented words are likely to be selected against. Starting from over- or underrepresented words corresponding to RMS recognition sites in specific clades, the specificity of unknown R-M systems can be highlighted. Among the known restriction enzymes, that are described in the REBASE database of restriction and modification systems, many of their recognition sites are still uncharacterized. Eventually, this motivates studies aimed at assessing horizontal transferring events of RMS in micro-organisms through the analysis of word usage biases in well-determined genomic regions. A probabilistic model is built on a first-order Markovian chain. Statistics on the k-neighborhood of a word is carried out to assess the biological significance of a genomic motif. Efficient word counting procedures have been implemented and statistics are used for the assessment of the significance of individual words in large sequences. On the basis of the set of most avoided words, and in accordance to the IUPAC coding standards, suggestions are made regarding potential recognition sequences. In certain cases, a comparison of avoided palindromic words in taxonomically related bacteria shows a pattern of relatedness of their R-M systems. For strengthening this analysis, the primary protein structure of all type II R-M systems known in REBASE have been blasted against the nr-GENBANK database. The combination of these analyses has revealed some interesting examples of possible horizontal transfer events of R-M systems.     

Mutagenic primer design for mismatch PCR-RFLP SNP genotyping using a genetic algorithm

Polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) is useful in small-scale basic research studies of complex genetic diseases that are associated with single nucleotide polymorphism (SNP). Designing a feasible primer pair is an important work before performing PCR-RFLP for SNP genotyping. However, in many cases, restriction enzymes to discriminate the target SNP resulting in the primer design is not applicable. A mutagenic primer is introduced to solve this problem. GA-based Mismatch PCR-RFLP Primers Design (GAMPD) provides a method that uses a genetic algorithm to search for optimal mutagenic primers and available restriction enzymes from REBASE. In order to improve the efficiency of the proposed method, a mutagenic matrix is employed to judge whether a hypothetical mutagenic primer can discriminate the target SNP by digestion with available restriction enzymes. The available restriction enzymes for the target SNP are mined by the updated core of SNP-RFLPing. GAMPD has been used to simulate the SNPs in the human SLC6A4 gene under different parameter settings and compared with SNP Cutter for mismatch PCR-RFLP primer design. The in silico simulation of the proposed GAMPD program showed that it designs mismatch PCR-RFLP primers. The GAMPD program is implemented in JAVA and is freely available at http://bio.kuas.edu.tw/gampd/.     

Categoric prediction of metal ion mechanisms in the active sites of 17 select type II restriction endonucleases

Recently determined crystal structures of type II restriction endonucleases have produced a plethora of information on the basis for target site sequence selectivity. The positioning and role of metal ions in DNA recognition sites might reflect important properties of protein-DNA interaction. Although acidic and basic groups in the active sites can be identified, and in some cases divalent-metal binding sites delineated, a convincing picture clarifying the way in which the attacking hydroxide ion is generated, and the leaving group stabilized, has not been elucidated for any of the enzymes. We have examined the interatomic distances between metal ions and proposed key catalytic residues in the binding sites of seventeen type II restriction endonucleases whose crystal structures are documented in literature. The summary and critical evaluation of structural assignments and predictions made earlier have been useful to group these enzymes. All the enzymes used for this study have been categorized on the basis of the number of metal ions identified in their crystal structures. Among 17 experimentally characterized (not putative) type II REases, whose apparently full-length sequences are available in REBASE, we predict 8 (47%) to follow the single metal ion mechanism, 5 to follow the two metal ion mechanism, 2, the three metal ion mechanism, 1, the four metal ion mechanism and 1 the six metal ion mechanism.     

Bioaccessibility and other key parameters in assessing oral exposure to PAH-contaminated soils and dust: A critical review

Soil ingestion is an important pathway for human exposure to polycyclic aromatic hydrocarbon (PAH)-contaminated soils and dust for children (via ingesting hand residue) as well as for adults (via occupational exposure). An appropriate selection of exposure parameter values is essential for having an accurate risk assessment. This review addresses key parameters for estimating oral exposure to PAH-contaminated soils/dust, discusses their variability and uncertainty, and provides recommendations for value selection. Bioaccessibility (contaminant fraction solubilized in gastro-intestinal tract, available for entering bloodstream and reaching target organs) and soil ingestion rate are two key parameters for exposure assessment (usually characterized by large variability and/or uncertainty), followed by exposure frequency/duration and body weight.     

BioCaster: detecting public health rumors with a Web-based text mining system

SUMMARY: BioCaster is an ontology-based text mining system for detecting and tracking the distribution of infectious disease outbreaks from linguistic signals on the Web. The system continuously analyzes documents reported from over 1700 RSS feeds, classifies them for topical relevance and plots them onto a Google map using geocoded information. The background knowledge for bridging the gap between Layman's terms and formal-coding systems is contained in the freely available BioCaster ontology which includes information in eight languages focused on the epidemiological role of pathogens as well as geographical locations with their latitudes/longitudes. The system consists of four main stages: topic classification, named entity recognition (NER), disease/location detection and event recognition. Higher order event analysis is used to detect more precisely specified warning signals that can then be notified to registered users via email alerts. Evaluation of the system for topic recognition and entity identification is conducted on a gold standard corpus of annotated news articles. AVAILABILITY: The BioCaster map and ontology are freely available via a web portal at http://www.biocaster.org.     

SNPicker: a graphical tool for primer picking in designing mutagenic endonuclease restriction assays

Simple, low-cost and accurate genotyping methods for single nucleotide polymorphisms (SNPs) are in high demand in the post-genome-sequencing era. We present a graphical tool called SNPicker, implemented in Java, which significantly facilitates the design of mutagenic endonuclease restriction assays. SNPicker uses the online NEB REBASE to automatically scan for all possible designs of mutagenic primers that can facilitate the picking of mismatched PCR primers to artificially introduce or abolish a restriction site at the target SNP site. We successfully applied SNPicker in designing endonuclease restriction assays for 14 SNPs for the MTHFR gene, the Coagulation Factor II gene and the Coagulation Factor V gene. The SNP assays designed using SNPicker were cross-validated using the MassARRAY technology.     

A spatio-temporal mining approach towards summarizing and analyzing protein folding trajectories

This paper presents a software library, nicknamed BATS, for some basic sequence analysis tasks. Namely, local alignments, via approximate string matching, and global alignments, via longest common subsequence and alignments with affine and concave gap cost functions. Moreover, it also supports filtering operations to select strings from a set and establish their statistical significance, via z-score computation. None of the algorithms is new, but although they are generally regarded as fundamental for sequence analysis, they have not been implemented in a single and consistent software package, as we do here. Therefore, our main contribution is to fill this gap between algorithmic theory and practice by providing an extensible and easy to use software library that includes algorithms for the mentioned string matching and alignment problems. The library consists of C/C++ library functions as well as Perl library functions. It can be interfaced with Bioperl and can also be used as a stand-alone system with a GUI. The software is available at http://www.math.unipa.it/~raffaele/BATS/ under the GNU GPL.     

Analysis of the Campylobacter jejuni Genome by SMRT DNA Sequencing Identifies Restriction-Modification Motifs

Campylobacter jejuni is a leading bacterial cause of human gastroenteritis. The goal of this study was to analyze the C. jejuni F38011 strain, recovered from an individual with severe enteritis, at a genomic and proteomic level to gain insight into microbial processes. The C. jejuni F38011 genome is comprised of 1,691,939 bp, with a mol.% (G+C) content of 30.5%. PacBio sequencing coupled with REBASE analysis was used to predict C. jejuni F38011 genomic sites and enzymes that may be involved in DNA restriction-modification. A total of five putative methylation motifs were identified as well as the C. jejuni enzymes that could be responsible for the modifications. Peptides corresponding to the deduced amino acid sequence of the C. jejuni enzymes were identified using proteomics. This work sets the stage for studies to dissect the precise functions of the C. jejuni putative restriction-modification enzymes. Taken together, the data generated in this study contributes to our knowledge of the genomic content, methylation profile, and encoding capacity of C. jejuni.     

An expeditious synthesis of 4-hydroxy-2E-nonenal (4-HNE), its dimethyl acetal and of related compounds

The facile one step synthesis of 4-hydroxy-2(E)-nonenal and its dimethyl acetal via a cross-metathesis reaction between commercially available octen-3-ol and acrolein or its dimethyl acetal is reported. The method was extended to the synthesis of C₆ and C₁₂ 4-hydroxy-2(E)-enals, their dimethyl acetal and of the 4-hydroxy-2(E)-nonenoic acid (4-HNA).     

GEOmetadb: powerful alternative search engine for the Gene Expression Omnibus

The NCBI Gene Expression Omnibus (GEO) represents the largest public repository of microarray data. However, finding data in GEO can be challenging. We have developed GEOmetadb in an attempt to make querying the GEO metadata both easier and more powerful. All GEO metadata records as well as the relationships between them are parsed and stored in a local MySQL database. A powerful, flexible web search interface with several convenient utilities provides query capabilities not available via NCBI tools. In addition, a Bioconductor package, GEOmetadb that utilizes a SQLite export of the entire GEOmetadb database is also available, rendering the entire GEO database accessible with full power of SQL-based queries from within R. AVAILABILITY: The web interface and SQLite databases available at http://gbnci.abcc.ncifcrf.gov/geo/. The Bioconductor package is available via the Bioconductor project. The corresponding MATLAB implementation is also available at the same website.