Microarrays of virus-specific oligonucleotides may provide a method of screening samples for the presence or absence of a large variety of viruses simultaneously. Influenza viruses are ideal for evaluating such microarrays because of their genetic and host diversity, and the availability of an extensive sequence database. A collection of 476 influenza virus-specific oligonucleotides was spotted onto glass slides as probes. Viral RNAs were reverse transcribed and amplified by PCR, and the products were labeled with cyanine dyes. The presence of viruses and their identities were determined by hybridization. The fluorescence intensities of oligonucleotide spots were highly reproducible within each slide and satisfactorily proportional between experiments. However, the intensities of probe spots completely complementary to target sequences varied from background to saturation. The variations did not correlate with base composition, nucleotide sequence, or internal secondary structures. Therefore, thresholds for determining whether hybridization to a spot should be judged as positive were assigned individually. Considering only positive spots from probes predicted to be monospecific for influenza virus species, subtype, host source, or gene segment, this method made correct identifications at the species, hemagglutinin subtype, and gene segment levels. Monospecific neuraminidase (NA) subtype probes were insufficiently diverse to allow confident NA subtype assignment. Incorporating positive spots from polyspecific probes into the identification scheme gave similar results. Overall, the results demonstrate the potential of microarray-based oligonucleotide hybridization for multiple virus detection.
VirOligo is a database of virus-specific oligonucleotides. The VirOligo database consists of two tables, Common data and Oligo data. The Oligo data table contains PCR primers and hybridization probes used for detection of viral nucleic acids and the Common data table contains the experimental conditions used in their detection. Each oligonucleotide entry contains links to PubMed, GenBank, NCBI Taxonomy databases and BLAST. As of July 2001, the VirOligo database contains a complete listing of oligonucleotides specific to viral agents associated with bovine respiratory disease that were published in English in peer-reviewed journals. The viruses are bovine herpes virus types 1, 3, 4 and 5, bovine viral diarrhea virus, bovine parainfluenza 3 virus, bovine respiratory syncytial virus, bovine adenovirus, bovine rhinovirus, bovine coronavirus, bovine reovirus, bovine enterovirus and alcelaphine herpesvirus-1. The VirOligo database is being expanded to other viruses and can be accessed through the Internet at http://viroligo.okstate.edu/.
There have been many reports suggesting that soluble oligomers of amyloid β (Aβ) are neurotoxins causing Alzheimer's disease (AD). Although inhibition of the soluble oligomerization of Aβ is considered to be effective in the treatment of AD, almost all peptide inhibitors have been designed from the β-sheet structure (H14-D23) of Aβ(1-42). To obtain more potent peptides than the known inhibitors of the soluble-oligomer formation of Aβ(1-42), we performed random screening by phage display. After fifth-round panning of a hepta-peptide library against soluble Aβ(1-42), novel peptides containing arginine residues were enriched. These peptides were found to suppress specifically 37/48 kDa oligomer formation and to keep the monomeric form of Aβ(1-42) even after 24 h of incubation, as disclosed by SDS-PAGE and size-exclusion chromatography. Thus we succeeded in acquiring novel efficient peptides for inhibition of soluble 37/48 kDa oligomer formation of Aβ(1-42).
Bridge Technology is an amplification technique in which pairs of primers are immobilized on a solid support, allowing amplification only at the location of the primer pair spot. The technique has diagnostic potential since an array of primer pairs, each specific for a different pathogen, can be used with a diagnostic sample without inter-pair interactions that plague the development of multiplex PCRs. As a result, one assay should be able to determine which of multiple pathogens are present and which are absent in each sample. As test material, we examined the specificity of detection of the RNA-containing bovine viral diarrhea virus (BVDV) and two DNA-containing bovine herpesviruses 1 and 2 (BHV-1 and BHV-2). Nylon membranes with two spots of UV-immobilized primer pairs--one for BVDV and one for BHV--were used in amplification with both corresponding templates, with each template singly and with no template. When amplification was assayed by chemiluminescent detection of incorporated DIG-nucleotides, the expected amplification patterns were obtained.
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