Abstract: The present invention demonstrates that gene expression can be controlled in vitro using DNA (gene) sequences immobilized on a template with micron scale temperature heaters. Such expression is controllable by varying temperature of the template on a short time scale. The present invention further demonstrates that nucleic acid constructs controlled by the present method express protein either free or bound to the nucleic acid. Based on these findings, the present invention provides methods and apparatuses useful for the preparation of in vitro programmable protein networks and protein micro arrays.

Abstract: The present invention relates to a purified or isolated polynucleotide encoding human CIDE B protein, the regulatory nucleic acids contained therein, polymorphic markers thereof, and the resulting encoded protein, as well as to methods and kits for detecting this polynucleotide and this protein. The present invention also pertains to a polynucleotide carrying the natural regulatory regions of the CIDE B gene which is useful, for example, to express a heterologous nucleic acid in host cells or host organisms as well as functionally active regulatory polynucleotides derived from said regulatory regions.

Abstract: Methods of genotyping amplified mixtures of DNAs, nucleic acid markers and methods of obtaining markers, kits, recombinant plants, positional cloning and integrated systems for making genotypes and assessing hybridizations are provided. These features are applicable to DNA fingerprinting, marker assisted selection, genotyping, cladistic analysis of variance, and high throughput laboratory screening methods.

Abstract: This invention provides a method of sorting genes comprising: (1) preparing ds cDNA molecules from mRNA molecules (2) digesting the ds cDNA molecules (3) ligating to the digested cDNA molecules a set of dsDNA oligonucleotide adaptors; (4) amplifying the ligated cDNA molecules; and (5) sorting the amplified cDNA molecules into nonoverlapping groups. This invention also provides two additional methods of sorting genes. This invention further provides a method of making sub-libraries of ligation sets.

Abstract: Methods of selecting tag nucleic acids and VLSIPS™ arrays and the arrays made by the methods are used to label and track compositions, including cells and viruses, e.g., in libraries of cells or viruses. In addition to providing a way of tracking compositions in mixtures, the tags facilitate analysis of cell and viral phenotypes.

Abstract: Methods and compositions to remove a nucleotide sequence of interest in a plant and plant cell are provided. In particular the methods of the invention comprise providing a plant cell having stably incorporated into its genome a transfer cassette comprising a nucleotide sequence of interest flanked by non-identical recombination sites and introducing into the plant cell a chimeric RNA-DNA oligonucleotide molecule. The chimeric RNA-DNA oligonucleotide is capable of recognizing and implementing a nucleotide conversion in one of the non-identical recombination sites so as to create two identical recombination sites. An appropriate recombinase is provided which excises the sequences between the two identical recombination sites.

Abstract: A process to obtain linear double-stranded covalently closed DNA “dumbbell” constructs from plasmids by restriction digest, subsequent ligation with hairpin oligodesoxyribonucleotides, optionally in the presence of restriction enzyme, and a final digestion with endo- and exonucleolytic enzymes that degrade all contaminating polymeric DNA molecules but the desired construct. The invention also provides a process to obtain said dumbbell constructs employing endonuclease class II enzymes. Furthermore, the invention provides a process to obtain linear, covalently closed DNA molecules, such as plasmids, free from contamination by genomic DNA, by submitting the DNA preparation to a facultative endonucleolytic degradation step and an obligatory exonucleolytic degradation step.

Abstract: A fluorescent cyanine dye of the following general formula is disclosed: wherein: X1 and X2 are independently selected from the group consisting of —O—, —S—, —C(CH3)2 or —C═CH2; Y1 and Y2 are nonmetal atoms required to form a benzo-condensed or naphtho-condensed ring; Q is a conjugated moiety that increases the fluorescent quantum yield and the stability of the compound; R1 and R2 are independently selected from the group consisting of H, C1-C4, alkyl, alkylensulfonic group or alkylensulfonate group wherein the alkylene group has from 1 to 4 carbon atoms; R3, R4 and R5 are independently selected from the group consisting of H, a sulfonic group, a sulfonate group, alkylensulfonic, alkylensulfonate and —SO2NH(CH2)m—W—(CH2)nZ, wherein alkylene has 1 to 4 carbon atoms, with the proviso that at least one of R1 to R5 contains a sulfonic or sulfonate group; W is absent or is a group selected from —SO2NH, —O—, —COO

Abstract: Specificity in immune responses is in part controlled by the selective interaction of T cell receptors with their cognate ligands, peptide/MHC molecules. The discriminating nature of this interaction makes these molecules, in soluble form, good candidates for selectively regulating immune responses. Attempts to exploit soluble analogs of these proteins has been hampered by the intrinsic low avidity of these molecules for their ligands. To increase the avidity of soluble analogs for their cognates to biologically relevant levels, divalent peptide/MHC complexes or T cell receptors (superdimers) were constructed. Using a recombinant DNA strategy, DNA encoding either the MHC class II/peptide or TCR heterodimers was ligated to DNA coding for murine Ig heavy and light chains. These constructs were subsequently expressed in a baculovirus expression system.

Abstract: The present invention relates to vanilloid receptor-2, a novel member of the vanilloid receptor family. The invention provides isolated nucleic acid molecules encoding human VR2 receptors. VR2 polypeptides are also provided, as are vectors, host cells and recombinant methods for producing the same. The invention further relates to screening methods for identifying agonists and antagonists of VR2 receptor activity. Also provided are diagnostic methods for detecting disease states related to the aberrant expression of VR2 receptors. Further provided are therapeutic methods for treating disease states including, but not limited to, chronic pain syndromes, congenital pain insensitivity, inflammation, ischemia, host defense dysfunction, immune surveillance dysfunction, arthritis, multiple sclerosis, autoimmunity, immune dysfunction, and allergy.

Abstract: The present invention relates to a method for the preparation of a protein by yeasts. In particular, the method of the invention concerns transforming a yeast cell with a first DNA fragment encoding the protein and a second DNA fragment that encodes the receptor. The method further entails culturing the transformed yeast cell, and isolating the protein. The first DNA fragment is under control of elements providing for expression of the DNA fragment in yeast, which elements include a higher eukaryotic positive transcription control sequence consisting of a natural ligand responsive element activating sequence or a variant. The receptor is a natural nuclear receptor selected from the group consisting of receptors for steroids or for retinoids or for thyroid hormones or for vitamin D3. The receptor includes a first fragment that recognizes said ligand and a second fragment that binds to said transcriptional control sequence.

Abstract: The invention relates to methods for identifying inhibitors of mucin production, methods for inhibiting mucin production and methods for treating airway diseases, such as cystic fibrosis, chronic bronchitis, bronchial pneumonia and asthma. Compositions are provided for use in the method comprising reporter gene constructs which are inducible by mucomones.

Abstract: Described herein are methods and reagents for encoding and sorting in vitro translated proteins.

Abstract: Clones containing a sequence encoding a glucuronide repressor are described. The nucleotide and amino acid sequences of a repressor (gusR) are presented. A glucuronide repressor is used to control expression of a transgene, detect glucuronides in a sample, and isolate glucuronides from a sample, among other uses.

Abstract: The invention provides new methods for modulating specific CMI-inducing cytokines in vivo. Such new methods result in stimulation of the cytokines IL-6, IL-12 MIP-1&bgr; and MCP without substantially inducing undesired cytokines. The methods according to the invention are based upon administration of oligonucleotides containing particular structural motifs which lead to specific cytokine induction.

Abstract: This invention provides methods for the discovery of molecules that target an essential aspect of eukaryotic gene expression—the formation of the mRNA 5′ cap m7GpppN. An underlying principle of this invention is the use of a different strains of a test organism that differ only in the composition or source of the essential cap-forming enzymes. The invention provides isogenic yeast strains that derive all their capping activities from fungal sources versus mammalian sources. These strains form the basis of a differential growth inhibition assay to identify molecules that specifically target the fungal capping apparatus. This invention also provides a method to screen in vitro for molecules that inhibit fungal RNA triphosphatase, an essential enzyme that catalyzes the first of three steps in cap synthesis.

Abstract: Modified dimers having a ribose sugar moiety in the 5′ nucleoside and a 2′ modified sugar in the 3′ nucleoside are provided. The modified dimers are useful in the preparation of oligonucleotide analogs having enhanced properties compared to native oligonucleotides, including increased nuclease resistance, enhanced binding affinity and improved protein binding.

Abstract: DNA fragments which contain a sequence of DNA which encodes a protective peptide-fused &agr;-hANP in which the protective peptide has a C-terminus lysine residue which is directly fused to the N-terminus of the &agr;-hANP, vectors which contain such a DNA sequence, and microorganisms transformed which such a vector are useful for the production of &agr;-hANP.

Abstract: The present invention is directed to the cloning, sequencing and expression of homologous immunoreactive 28-kDa protein genes, ECa28-1 and ECa28SA3, from a polymorphic multiple gene family of Ehrlichia canis. A complete sequence of another 28-kDa protein gene, ECaSA2, is also provided. Further disclosed is a multigene locus encoding all five homologous 28-kDa protein genes of Ehrlichia canis. Recombinant Ehrlichia canis 28-kDa proteins react with convalescent phase antiserum from an E. canis-infected dog.

Abstract: The invention relates to methods for the sensitive, specific, and, preferably, quantitative detection of C. parvum oocysts in aqueous samples.

Abstract: The present invention is directed to an approach to identify changes in gene expression by selective amplification of 3′ fragments of double stranded cDNAs.

Abstract: The present invention provides novel identification polypeptides containing multiple copies of an antigenic domain joined in tandem to provide increased sensitivity for the detection and purification of target peptides, a cleavable linking sequence and optionally a spacer domain. Further provided are hybrid polypeptide molecules composed of an identification polypeptide and a target peptide which are produced by recombinant DNA technology and purified using affinity chromatography using one or more ligands. Accordingly, also provided are DNA expression vectors containing DNA encoding for identification polypeptides and methods for using such identification polypeptides for the purification of target peptides. Also provided are methods of constructing DNA vectors encoding the novel identification polypeptides and DNA expression vectors encoding the identification polypeptides linked to a target peptide.

Abstract: A molecule comprising a nucleic acid portion and a protein portion directly bound to said nucleic acid portion with a covalent bond, wherein said nucleic acid portion comprises a polymer of nucleoside, and said protein portion is encoded by said nucleic acid portion, and a method for constructing the molecule, which comprises (a) preparing a DNA containing a gene which has no termination codon, (b) transcribing the prepared DNA into RNA, (c) bonding a chimeric spacer composed of DNA and RNA to a 3′-terminal end of the obtained RNA, (d) bonding to a 3′-terminal end of the obtained bonded product, a nucleoside or a substance having a chemical structure analogous to that of a nucleoside, which can be covalently bound to an amino acid or substance having a chemical structure analogous to that of an amino acid, and (e) performing protein synthesis in a cell-free protein synthesis system using the obtained bonded product as mRNA to bond a nucleic acid portion containing the gene to a translation product

Abstract: Methods are disclosed for identifying an RNA fragment that mimics the structure of a defined or undefined target RNA molecule to which a compound binds inside of a cell resulting in retardation of cell growth or cell death. Methods using these RNA fragments for identifying unknown compounds of pharmaceutical interest, and for identifying unknown RNA targets for use in treating disease are disclosed. These methods and compositions are used in screening for novel antibiotics, bacteriostatics, or modifications thereof or for identifying compounds useful to alter expression levels of proteins encoded by mRNA. The methods involve providing random DNA fragments from DNA which encodes RNA target molecules, cloning such fragments to create a plasmid library of same; transfecting cells which contain the native RNA target molecule with the plasmid library and exposing the cells to one or more of test compounds.

Abstract: Transcription promoter and terminator sequences from the Pichia methanolica glyceraldehyde-3-phosphate dehydrogenase 2 gene (GAP2 gene) are disclosed. The sequences are useful within DNA constructs for the production of proteins of interest in cultured P. methanolica cells. Within the expression vectors, a GAP2 promoter and/or a GAP2 terminator is operably linked to a DNA segment encoding the protein of interest.

Abstract: A new mumps vaccine is presented, comprising a homogeneous pure isolate derived from the Jeryl-Lynn strain of mumps virus.

Abstract: The present invention describes an efficient retroviral or viral based method that allows easy and quick identification of gene transfer in living, transduced mammalian cells. Retroviral and viral vector producer cells were generated containing a gene for an improved humanized red-shifted, Green Fluorescent Protein (hRGFP) which increases the resulting fluorescence yield after excitation. This humanized, red-shifted GFP (hRGFP) gene was cloned into several vectors and transfected into various packaging cell lines to produce vibrant green fluorescence after excitation with blue light at 450-490 nm. These vectors represent a substantial advance over currently available gene transfer marking systems or wild-type GFP marker systems none of which have been stably transfected into cells.

Abstract: The present invention provides methods and compositions for preparations of recombinant parvovirus virions with a reduced number of replication competent particles. The compositions of the present invention include nucleic acids encoding parvovirus helper functions which contain at least one non-native intron sequence. The present invention also includes helper function vectors, host cells transfected with the helper function vectors, methods of using the helper function vectors, and recombinant parvovirus virions produced by such methods.

Abstract: The present invention provides a gene, termed “FT” for flowering locus T, and a polypeptide encoded by FT that modulates flower development in plants. FT is useful in methods of the invention for producing genetically modified plants characterized as having the phenotypic trait of modulated flower development, for example early or delayed flowering. Such plants can be genetically modified by nucleic acids encoding functional FT peptides; at least one antisense nucleic acid for FT; a structural gene that encodes wild-type FT polypeptide; or a structural gene that encodes dominant negative polypeptides, for example, in order to modulate flowering in the plant.

Abstract: A method of preparing an immobilized oligonucleotide having a free 3′-end comprises the steps of: i) preparing an oligonucleotide attached in a first position to a solid support via its 3′-end and having a free 5′-end; ii) binding said oligonucleotide in a second position remote from the 3′-end to the solid support; and iii) selectively releasing the 3′-end of the oligonucleotide from the solid support to obtain the oligonucleotide attached to the support in said second position in a reversed orientation with a free 3′-end.

Abstract: An expression system for producing and isolating large quantities of protein. The system comprises an expression vector containing a first coding region which codes for glutathione-S-transferase operatively connected to a baculovirus promoter, a second coding region in-frame with the first coding region, and a restriction region downstream of the first coding region, into which the second coding region is inserted. A fusion protein encoded by the first and second coding region is produced by expression of the vector. Examples of this second coding region include Lck, LynB, Syk, Blk, Fyn, and Yes. A process for expression of the vector in a host cell such as Spodoptera frugiperda is also included.

Abstract: The present invention generally provides a rapid efficient method for analyzing polymorphic or biallelic markers, and arrays for carrying out these analyses. In general, the methods of the present invention employ arrays of oligonucleotide probes that are complementary to target nucleic acids which correspond to the marker sequences of an individual. The probes are typically arranged in detection blocks, each block being capable of discriminating the three genotypes for a given marker, e.g., the heterozygote or either of the two homozygotes. The method allows for rapid, automatable analysis of genetic linkage to even complex polygenic traits.

Abstract: The invention relates to a method for analyzing of determining the methylation pattern of a starting DNA and/or for distinguishing between methylated and non-methylated sites in the starting DNA, comprising at least (A) generating a first DNA fingerprint, containing bands corresponding to both the methylated and non-methylated sites of interest; and/or (B) generating a second DNA fingerprint, containing bands corresponding only to the methylated sites of interest; and optionally comprising (C) generating a third DNA fingerprint, containing bands corresponding only to the non-methylated sites of interest; and optionally further comprising (D) analysing the fingerprint(s) thus obtained. The fingerprints are preferably generated using AFLP, by means of a frequent cutter and a methylation sensitive rare cutter. The invention further relates to specific methods for generating the above first and second DNA fingerprint by means of AFLP, and kits for use with said methods.

Abstract: The present invention relates to novel methods for sequencing and mapping genetic markers in polynucleotide sequences using Type-IIs restriction endonucleases. The methods herein described result in the “capturing” and determination of specific oligonucleotide sequences located adjacent to Type-IIs restriction sites. The resulting sequences are useful as effective markers for use in genetic mapping, screening and manipulation.

Abstract: This invention describes a nucleic acid delivery vehicle construct for transfecting and/or infecting a target cell. The construct is made of a delivery vehicle and a bifunctional complex for linking the delivery vehicle to a target cell. The bifunctional complex has a delivery vehicle-binding molecule or fragment (“delivery vehicle-binding portion”), a molecule or fragment thereof that binds to a cell surface molecule on the target cell (“cell surface molecule-binding portion”) and a linker which connects the molecules or fragments.

Abstract: The present invention relates to a novel protein, the Human Oncogene Induced Secreted Protein I (“HOIPS I”) protein. In particular, isolated nucleic acid molecules are provided encoding the human HOIPS I protein. HOIPS I polypeptides are also provided as are vectors, host cells and recombinant methods for producing the same. Also provided are diagnostic methods for detecting abnormal cell proliferation and differentiation disorders and therapeutic methods for treating the same.

Abstract: A restriction site indexing method for selectively amplifying any fragment generated by a Class II restriction enzyme includes adaptors specific to fragment ends containing adaptor indexing sequences complementary to fragment indexing sequences near the termini of fragments generated by Class II enzyme cleavage A method for combinatorial indexing facilitates amplification of restriction fragments whose sequence is not known. Profiling methods and other methods for characterizing polynucleotides are presented.

Abstract: Described herein are methods and reagents for the selection of protein molecules that make use of RNA-protein fusions.

Abstract: Recombinational cloning is provided by the use of nucleic acids, vectors and methods, in vitro and in vivo, for moving or exchanging segments of DNA molecules using engineered recombination sites and recombination proteins to provide chimeric DNA molecules that have the desired characteristic(s) and/or DNA segment(s).

Abstract: A circular DNA molecule, useful for gene therapy, comprising at least one nucleic acid sequence of interest, characterized in that the region allowing the replication thereof has an origin of replication with a functionality in a host cell that requires the presence of at least one specific protein foreign to said host cell. A method for preparing same, cells incorporating said DNA molecules and uses thereof in gene therapy are also described.

Abstract: A support for solid phase amplification or sequencing of nucleic acids has a functionalized solid support, a linker arm having functional groups covalently bound to the solid support through at least one binding site of the functional groups, and an oligonucleotide primer bound at its 5′ end to the linker, the primer, thereby, being immobilized on the solid support.

Abstract: A method for detecting gene expression in cells by reverse transcribing mRNA molecules into cDNA, cutting the cDNA with at least one restriction endonuclease, adding adaptor sequences to the cDNA fragments and selectively amplifying a subset of the cDNA by a polymerase chain reaction (PCR) to present a two-dimensional display of the DNA fragments or for cloning the DNA fragments into a vector is disclosed. In one embodiment, cDNA corresponding to the 3′ end of the mRNA is amplified and displayed or cloned, whereas in another embodiment, cDNA corresponding to the entire mRNA molecule is amplified and displayed or cloned.

Abstract: The invention is a method of detecting nucleic acids in a sample using oligonucleotide probes which are noncovalently bound to solid supports for rapid, sensitive, hybridization assays. The method involves coating the support surface with a polynucleotide and then hybridizing a specific capture probe for each analyte to the polynucleotide by way of a short tail of the complementary polynucleotide. The immobilized probes are used to capture nucleic acid targets out of complex specimens for nonisotopic detection without the need for prior cell culture or purification of the target nucleic acids. A panel of tests can be run on each specimen simultaneously, a format that conserves precious samples. The assay can be readily automated, and can be conveniently run in a manual fashion on large numbers of samples in two to three hours.

Abstract: Disclosed is a method for the comprehensive analysis of nucleic acid samples and a detector composition for use in the method. The method, referred to as Fixed Address Analysis of Sequence Tags (FAAST), involves generation of a set of nucleic acid fragments having a variety of sticky end sequences; indexing of the fragments into sets based on the sequence of sticky ends; associating a detector sequence with the fragments; sequence-based capture of the indexed fragments on a detector array; and detection of the fragment labels. Generation of the multiple sticky end sequences is accomplished by incubating the nucleic acid sample with one or more nucleic acid cleaving reagents. The indexed fragments are captured by hybridization and coupling, preferably by ligation, to a probe. The method allows a complex sample of nucleic acid to be quickly and easily cataloged in a reproducible and sequence-specific manner.

Abstract: The invention relates to a recombinant poliovirus vector and method of making the vector. The recombinant poliovirus vector comprises an RNA polymerase promoter operably linked to a poliovirus nucleic acid sequence which has been altered by removal of the CelII-SnaBI fragment and insertion of a heterologous nucleic acid sequence encoding a protein of interest. The heterologous nucleic acid sequence is flanked on both sides by internal ribosomal entry sites which allow for the expression of the encoded protein of interest and subsequent proteins encoded by the altered poliovirus nucleic acid sequence.

Abstract: Described herein is a method for selectively inhibiting the amplification of a specific DNA template during a polymerase chain reaction (PCR). In particular, the method is useful when the sequences of the desired and undesired DNA templates are similar. A set of universal primers binds to both the desired and undesired DNA templates during a PCR, resulting in the amplification of their DNA sequences. The method targets the undesired DNA template with three sets of oligonucleotide primers, one set of which is terminally modified to both prevent primer extension and increase the primer-template binding affinity. The result of these terminal modifications is the specific inhibition of the PCR amplification of the undesired DNA template, allowing the preferential amplification of the desired DNA templates.

Abstract: Methods and kits for labeling nucleic acids are provided. In the subject methods, an oligonucleotide tagged nucleic acid comprising an oligonucleotide tag is first generated. The oligonucleotide tagged nucleic acid is then contacted under hybridization conditions with a labeled oligonucleotide complementary to the oligonucleotide tag, yielding a labeled nucleic acid. The kits of the subject invention at least include a primer for use in enzymatically generating an oligonucleotide tagged target nucleic acid, where the primer generally at least includes an oligo dT region and the oligonucleotide tag, and a labeled oligonucleotide complementary to the oligonucleotide tag. The subject methods and kits find use in a variety of applications, and are particularly suited for use in gene expression analysis applications.

Abstract: A method is disclosed for detecting the presence of a difference between two related nucleic acid sequences. In the method a complex is formed comprising both strands of each sequence. Each member of at least one pair of non-complementary strands within the complex have labels. The association of the labels as part of the complex is determined as an indication of the presence of a difference between the two related sequences. The complex generally comprises a Holliday junction. In one aspect a medium suspected of containing said two related nucleic acid sequences is treated to provide partial duplexes having non-complementary tailed portions at one end. The double stranded portions of the partial duplexes are identical except for said difference.

Abstract: Recombinant tropoelastins and variants of recombinant tropoelastins produced from synthetic polynucleotides, as well as the synthetic polynucleotides themselves are provided. Also provided are cross-linked elastins or elastin-like products prepared from the tropoelastins or variants.

Abstract: A yeast &agr;-factor expression system is provided comprised of a truncated leader sequence, containing the &agr;-factor signal peptide and one glycosylation site, linked by a processing site to a non-yeast protein-encoding sequence.