Enhanced SPARQL-based design rationale retrieval

Design rationale (DR) is an important category within design knowledge, and effective reuse of it depends on its successful retrieval. In this paper, an ontology-based DR retrieval approach is presented, which allows users to search by entering normal queries such as questions in natural language. First, an ontology-based semantic model of DR is developed based on the extended issue-based information system-based DR representation in order to effectively utilize the semantics embedded in DR, and a database of ontology-based DR is constructed, which supports SPARQL queries. Second, two SPARQL query generation methods are proposed. The first method generates initial SPARQL queries from natural language queries automatically using template matching, and the other generates initial SPARQL queries automatically from DR record-based queries. In addition, keyword extension and optimization is conducted to enhance the SPARQL-based retrieval. Third, a design rationale retrieval prototype system is implemented. The experimental results show the advantages of the proposed approach

miRecords: an integrated resource for microRNA–target interactions

MicroRNAs (miRNAs) are an important class of small noncoding RNAs capable of regulating other genes’ expression. Much progress has been made in computational target prediction of miRNAs in recent years. More than 10 miRNA target prediction programs have been established, yet, the prediction of animal miRNA targets remains a challenging task. We have developed miRecords, an integrated resource for animal miRNA–target interactions. The Validated Targets component of this resource hosts a large, high-quality manually curated database of experimentally validated miRNA–target interactions with systematic documentation of experimental support for each interaction. The current release of this database includes 1135 records of validated miRNA–target interactions between 301 miRNAs and 902 target genes in seven animal species. The Predicted Targets component of miRecords stores predicted miRNA targets produced by 11 established miRNA target prediction programs. miRecords is expected to serve as a useful resource not only for experimental miRNA researchers, but also for informatics scientists developing the next-generation miRNA target prediction programs. The miRecords is available at http://miRecords.umn.edu/miRecords

NMR structure and Mg(2+) binding of an RNA segment that underlies the L7/L12 stalk in the E.coli 50S ribosomal subunit

Helix 42 of Domain II of Escherichia coli 23S ribosomal RNA underlies the L7/L12 stalk in the ribosome and may be significant in positioning this feature relative to the rest of the 50S ribosomal subunit. Unlike the Haloarcula marismortui and Deinococcus radiodurans examples, the lower portion of helix 42 in E.coli contains two consecutive G•A oppositions with both adenines on the same side of the stem. Herein, the structure of an analog of positions 1037–1043 and 1112–1118 in the helix 42 region is reported. NMR spectra and structure calculations support a cis Watson–Crick/Watson–Crick (cis W.C.) G•A conformation for the tandem (G•A)(2) in the analog and a minimally perturbed helical duplex stem. Mg(2+) titration studies imply that the cis W.C. geometry of the tandem (G•A)(2) probably allows O6 of G20 and N1 of A4 to coordinate with a Mg(2+) ion as indicated by the largest chemical shift changes associated with the imino group of G20 and the H8 of G20 and A4. A cross-strand bridging Mg(2+) coordination has also been found in a different sequence context in the crystal structure of H.marismortui 23S rRNA, and therefore it may be a rare but general motif in Mg(2+) coordination

Accurate multiplex gene synthesis from programmable DNA microchips

Testing the many hypotheses from genomics and systems biology experiments demands accurate and cost-effective gene and genome synthesis. Here we describe a microchip-based technology for multiplex gene synthesis. Pools of thousands of 'construction' oligonucleotides and tagged complementary 'selection' oligonucleotides are synthesized on photo-programmable microfluidic chips(1), released, amplified and selected by hybridization to reduce synthesis errors ninefold. A one-step polymerase assembly multiplexing reaction assembles these into multiple genes. This technology enabled us to synthesize all 21 genes that encode the proteins of the Escherichia coli 30S ribosomal subunit, and to optimize their translation efficiency in vitro through alteration of codon bias. This is a significant step towards the synthesis of ribosomes in vitro and should have utility for synthetic biology in general.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/62677/1/nature03151.pd

High density peptide microarrays. In situ synthesis and applications

The technologies enabling the creation of large scale, miniaturized peptide or protein microarrays are emerging. The focuses of this review are the synthesis and applications of peptide and peptidomimetic microarrays, especially the light directed parallel synthesis of individually addressable high density peptide microarrays using a novel photogenerated reagent chemistry and digital photolithography (Gao et al., 1998, J. Am. Chem. Soc. 120, 12698; Pellois et al. 2002, Nat. Biotechnol. 20, 922). Concepts related to the synthesis are discussed, such as the reactions of photogenerated acids in the deprotection step of peptide synthesis or oligonucleotide synthesis, and the applications of high density peptide chips in antibody binding assays are discussed. Peptide chips provide versatile tools for probing antigen-antibody, protein-protein, peptide-ligand interactions and are basic components for miniaturization, automation, and system integration in research and clinical diagnosis applications.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/43247/1/11030_2004_Article_5263634.pd

Nucleotide modification at the γ-phosphate leads to the improved fidelity of HIV-1 reverse transcriptase

The mechanism by which HIV-1 reverse transcriptase (HIV-RT) discriminates between the correct and incorrect nucleotide is not clearly understood. Chemically modified nucleotides containing 1-aminonaphthalene-5-sulfonate (ANS) attached to their γ-phosphate were synthesized and used to probe nucleotide selection by this error prone polymerase. Primer extension reactions provide direct evidence that the polymerase is able to incorporate the gamma-modified nucleotides. Forward mutation assays reveal a 6-fold reduction in the mutational frequency with the modified nucleotides, and specific base substitutions are dramatically reduced or eliminated. Molecular modeling illustrates potential interactions between critical residues within the polymerase active site and the modified nucleotides. Our data demonstrate that the fidelity of reverse transcriptase is improved using modified nucleotides, and we suggest that specific modifications to the γ-phosphate may be useful in designing new antiviral therapeutics or, more generally, as a tool for defining the structural role that the polymerase active site has on nucleotide selectivity

A versatile microreactor platform featuring a chemical-resistant microvalve array for addressable multiplex syntheses and assays

A versatile microreactor platform featuring a novel chemical-resistant microvalve array has been developed using combined silicon/polymer micromachining and a special polymer membrane transfer process. The basic valve unit in the array has a typical ‘transistor’ structure and a PDMS/parylene double-layer valve membrane. A robust multiplexing algorithm is also proposed for individual addressing of a large array using a minimal number of signal inputs. The in-channel microvalve is leakproof upon pneumatic actuation. In open status it introduces small impedance to the fluidic flow, and allows a significantly larger dynamic range of flow rates (∼ml min−1) compared with most of the microvalves reported. Equivalent electronic circuits were established by modeling the microvalves as PMOS transistors and the fluidic channels as simple resistors to provide theoretical prediction of the device fluidic behavior. The presented microvalve/reactor array showed excellent chemical compatibility in the tests with several typical aggressive chemicals including those seriously degrading PDMS-based microfluidic devices. Combined with the multiplexing strategy, this versatile array platform can find a variety of lab-on-a-chip applications such as addressable multiplex biochemical synthesis/assays, and is particularly suitable for those requiring tough chemicals, large flow rates and/or high-throughput parallel processing. As an example, the device performance was examined through the addressed synthesis of 30-mer DNA oligonucleotides followed by sequence validation using on-chip hybridization. The results showed leakage-free valve array addressing and proper synthesis in target reactors, as well as uniform flow distribution and excellent regional reaction selectivity.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/49051/2/jmm6_8_001.pd