Abstract: This invention relates to agents and conjugates that can be used to detect and isolate target components from complex mixtures such as nucleic acids from biological samples, cells from bodily fluids, and nascent proteins from translation reactions. Agents comprise a detectable moiety bound to a photoreactive moiety. Conjugates comprise agents coupled to substrates by covalent bounds which can be selectively cleaved with the administration of electromagnetic radiation. Targets substances labeled with detectable molecules can be easily identified and separated from a heterologous mixture of substances. Exposure of the conjugate to radiation releases the target in a functional form and completely unaltered. Using photocleavable molecular precursors as the conjugates, label can be incorporated into macromolecules, the nascent macromolecules isolated and the label completely removed.

Abstract: Disclosed are a method, device kit, and automated system for simple, reproducible, and high-throughput quantification of mRNA from whole blood. More particularly, the method, device, kit and automated system involve combinations of leukocyte filters attached to oligo(dT)-immobilized multi-well plates.

Abstract: The present invention provides cold shock protein-containing compositions for improved DNA synthesis reactions with improved reactivity, methods for synthesizing DNA using such compositions, kits for use in such methods, and DNA compositions yielded by such methods. The present invention further provides cold shock protein-containing compositions for the identification of endoribonuclease cleavage sites, methods for identifying endoribonuclease cleavage sites using such compositions, and kits for use in such methods.

Abstract: This invention relates to the use of tumor-derived or associated extracellular ribonucleic acid (RNA) found circulating in the plasma or serum fraction of blood for the detection, monitoring, or evaluation of cancer or premalignant conditions. Specifically, this invention enables the extraction of circulating RNA from plasma or serum and utilizes nucleic acid amplification assays for the identification, detection, inference, monitoring, or evaluation of any neoplasm, benign, premalignant, or malignant, in humans or other animals, which might be associated with that RNA. Further, this invention allows the qualitative or quantitative detection of tumor-derived or associated extracellular RNA circulating in the plasma or serum of humans or animals with or without any prior knowledge of the presence of cancer or premalignant tissue.

Abstract: This invention relates to a composition, kit, or DNA chip comprising polynucleotides and antibodies as probes for detecting, determining, or predicting the presence or metastasis of esophageal cancer, and to a method for detecting, determining, or predicting the presence or metastasis of esophageal cancer using the same.

Abstract: The present invention provides methods for the amplification of nucleic acid molecules. Methods for amplifying target polynucleotides, including mRNA, using oligonucleotides, DNA and RNA polymerases are provided. The invention further provides compositions and kits for practicing the methods, as well as methods which use the amplification products.

Abstract: A process for assessing mitochondrial toxicity of a compound that includes contacting nucleic acids from a host with an amplification reaction mixture that contains at least two primers that provide detectable signals, wherein: a first primer provides a first detectable signal upon amplification of a host mitochondrial nucleic acid; a second primer provides a second detectable signal upon amplification of a host nuclear nucleic acid; and comparing the first and second detectable signals.

Abstract: Aspects of the disclosure are generally directed to probes and probe compositions for detecting or quantifying a target. One aspect provides a method for selectively hybridizing a probe to a polynucleotide by contacting a sample containing a first and second polynucleotide with a probe. The probe includes a number of nucleotides complementary to the first or second polynucleotide in the region of mismatch between the first and second polynucleotides. Another aspect provides arrays including the disclosed probes and methods of using the arrays and the probes.

Abstract: The current inventors have discovered that the incorporation of a triplex forming monomer unit into oligonucleotides surprisingly gives the oligonucleotide a number of favorable characteristics. The oligonucleotides are advantageous because they allow modulation of the melting temperature of an oligonucleotide, they have improved sequence specificity and they can form triplexes by Hoogsteen or reverse Hoogsteen base pairing with double stranded nucleic acids. Moreover, some of the oligonucleotides of the invention have useful fluorescent characteristics, and the oligonucleotides comprising a triplex forming monomer can be used as substrates for enzymatic manipulations such as primer extension.

Abstract: The present invention provides a new procedure for the synthesis of cDNA from single cells after microdissection. It has the advantage that it is cost-efficient and can be carried out quickly with only few steps, even by less skilled laboratory employees. For the first time, the time-consuming and risky step of RNA isolation is omitted during cDNA synthesis from single cells by performing lysis and cDNA synthesis in the same reaction tube and in one buffer solution, which provides reliable and contamination-free results. The buffer is composed of NP40, carrier-RNA and Super RNAsin, as well as dNTPs and cDNA synthesis primers.

Abstract: The invention describes a method for isolating one or more genetic elements encoding a gene product having a desired activity, comprising the steps of: (a) compartmentalising genetic elements into microcapsules; (b) expressing the genetic elements to produce their respective gene products within the microcapsules; (c) sorting the genetic elements which produce the gene product having the desired activity using a change in the optical properties of the genetic elements. The invention enables the in vitro evolution of nucleic acids and proteins by repeated mutagenesis and iterative applications of the method of the invention.

Abstract: The present methods are exemplified by a process in which maternal blood containing fetal DNA is diluted to a nominal value of approximately 0.5 genome equivalent of DNA per reaction sample. Digital PCR is then be used to detect aneuploidy, such as the trisomy that causes Down Syndrome. Since aneuploidies do not present a mutational change in sequence, and are merely a change in the number of chromosomes, it has not been possible to detect them in a fetus without resorting to invasive techniques such as amniocentesis or chorionic villi sampling. Digital amplification allows the detection of aneuploidy using massively parallel amplification and detection methods, examining, e.g., 10,000 genome equivalents.

Abstract: The invention relates to a method for synthesizing a nucleic acid containing modified nucleotides. The method encompasses the following steps: a matrix strand is provided; —a primer which at least partially hybridizes on the matrix strand is provided; —nucleoside triphosphates, at least some of which are modified nucleoside triphosphates, are provided; —a polymerase activity is supplied; and—the matrix strand, the primer, and the nucleoside triphosphates are incubated so as to synthesize a nucleic acid that is substantially complementary to the matrix strand. The polymerase activity can be a reverse transcriptase activity.

Abstract: This invention provides methods for assessing HPV infection. Gene expression levels are used to assess the progression of HPV infection from benign to malignant growth. Also provided are kits for carrying out the methods of this invention.

Abstract: The invention relates to a method for generating a DNA sequence coding for the heavy chain or the light chain of at least one antibody from RNA from a cell capable of producing an antibody. More particularly, the invention relates to the generation of a monoclonal antibody library. The invention also relates to the use of an antibody library for screening monoclonal antibodies, preferably human antibodies for treating cancer.

Abstract: The present invention provides a method of covalently joining a DNA strand to an RNA strand comprising (a) forming a topoisomerase-DNA intermediate by incubating a DNA cleavage substrate comprising a topoisomerase cleavage site with a topoisomerase specific for that site, wherein the topoisomerase-DNA intermediate has one or more 5? single-strand tails; and (b) adding to the topoisomerase-DNA intermediate an acceptor RNA strand complementary to the 5? single-strand tail under conditions permitting a ligation of the covalently bound DNA strand of the topoisomerase-DNA intermediate to the RNA acceptor strand and dissociation of the topoisomerase, thereby covalently joining the DNA strand to the RNA strand. The present invention also provides a method of tagging a 5? end of an RNA molecule. The present invention further provides a DNA-RNA molecule which has been joined in vitro by the use of a topoisomerase. The present invention also provides a method of tagging a 5? end of an mRNA.

Abstract: The present teachings provide methods, compositions, and kits for performing primer extension reactions. In some embodiments, a reverse transcription reaction is performed on a target polynucleotide with a hot start primer comprising a blunt-ended self-complementary stem, and a loop, and extension products form at high temperatures but reduce extension product formation at low temperatures.

Abstract: The present invention comprises novel methods, compositions and kits that use N4 vRNAP deletion mutants to detect and quantify analytes comprising one or multiple target nucleic acid sequences, including target sequences that differ by as little as one nucleotide or non-nucleic acid analytes, by detecting a target sequence tag that is joined to an analyte-binding substance. The method consists of an annealing process, a DNA ligation process, an optional DNA polymerase extension process, a transcription process, and, optionally, a detection process.

Abstract: Thermostable viral and microbial polymerases exhibiting a combination of activities selected from proofreading (3?-5?) exonuclease activity, nick translating (5?-3?) nuclease activity, synthetic primer-initiated polymerase activity, nick-initiated polymerase activity, reverse transcriptase activity, strand displacement activity, terminal transferase activity, primase activity, and/or efficient incorporation of chain terminating analogs. Some of the polymerases provided herein include a first motif and a second motif. The first motif preferably has the sequence X1X2X3DX4PX5IELRX6X7X8, wherein X1 is I or V; X4 is F or Y; X8 is G or A; and X2, X3, X5, X6, and X7 are any amino acid. The second motif preferably has the sequence RX9X10X11KSANX12GX13X14YG, wherein X11 is G or A; X12 is F, L, or Y; X13 is L or V; X14 is I or L; and X9 and X10 are any amino acid.

Abstract: This invention provides novel compositions and processes for analyte detection, quantification and amplification. Nucleic acid arrays and libraries of analytes are usefully incorporated into such compositions and processes. Universal detection elements, signaling entities and the like are employed to detect and if necessary or desirable, to quantify analytes. Amplification of target analytes are also provided by the compositions and processes of this invention.

Abstract: The present invention provides methods for analyzing exon expression profiles of a cell or type of cell. In the invention, the expression levels of a plurality of individual exons or multiexons for each of a plurality of genes in the genome of an organism are measured and analyzed to determine the biological state, such as the exon expression state or transcriptional state, of the cell or type of cell. The methods of the invention are useful for determination of alternative RNA splicing in a plurality of genes. The invention also provides nucleic acid probe arrays for determining in parallel the expression levels of a plurality of exons or multiexons for each of a plurality of genes in the genome of an organism. The invention further provides methods for determining the effects of perturbations, such as perturbations by drugs, on exon expression and alternative RNA splicing pathways.

Abstract: The present invention concerns a method for diagnosing or monitoring elevated intraocular pressure or glaucoma in a patient comprising providing a biological sample from the patient and measuring the expression level of a superoxide dismutase-2 (SOD-2) gene in the biological sample. The present invention also concerns methods for treating glaucoma or elevated intraocular pressure in a patient comprising administering to a patient an effective amount of an agent that modulates SOD-2 expression or SOD-2 activity.

Abstract: New RNA cap analogs are disclosed containing one or more phosphorothioates groups. The analogs also contain modifications at the 2?-O position of 7-methylguanosine that prevent them from being incorporated in the reverse orientation during in vitro synthesis of mRNA and that hence are “anti-reverse cap analogs” (ARCAs). The ARCA modification ensures that the S atom is precisely positioned within the active sites of cap-binding proteins in both the translational and decapping machinery. The new S-ARCA analogs are resistant to in vivo decapping enzymes. Some S-ARCAs have a higher affinity for eIF4E than the corresponding analogs not containing a phosphorothioate group. When mRNAs containing the various S-ARCAs are introduced into cultured cells, some are translated as much as five-fold more efficiently than mRNAs synthesized with the conventional analog m7GpppG.

Abstract: Compositions, methods and kits for detecting viral nucleic acids. Targets that can be detected in accordance with the invention include HBV and/or HIV-1 and/or HCV nucleic acids. Particularly described are oligonucleotides that are useful as hybridization probes and amplification primers that facilitate detection of very low levels of HBV nucleic acids.

Abstract: The present invention provides methods for identifying druggable targets in assays that feature compositions, cells and/or organisms having structured viral non-coding RNAs (svRNAs) and an RNA interference (RNAi) pathway. Methods for identifying antiviral agents and creating vaccines are also featured. The invention further provides methods for inhibiting RNAi involving svRNAs or inhibitory derivatives thereof. The invention also provides compositions for delivering siRNA and miRNA molecules derived from svRNA loci and methods of use thereof. Therapeutic methods are also featured.

Abstract: This invention provides methods for detecting the presence of malignant or premalignant cells, or trophoblastic cells in a human wherein the malignant, premalignant or trophoblastic cells express 5T4. The methods of the invention detect 5T4 RNA in blood, blood plasma, serum, and other bodily fluids. The inventive methods are useful for detection, diagnosis, monitoring, treatment, or evaluation of neoplastic disease, and for the detection and evaluation of placental tissue in pregnant women.

Abstract: Disclosed are a method, device kit, and automated system for simple, reproducible, and high-throughput quantification of mRNA from whole blood. More particularly, the method, device, kit and automated system involve combinations of leukocyte filters attached to oligo(dT)-immobilized multi-well plates.

Abstract: Methods and kits for selective preparing cDNA relatively free of sequences found in rRNA and subcellular RNAs are disclosed. The methods and kits utilize approximately 200 hexamer sequences which target messenger RNA. The methods and kits are useful in preparing samples for sequencing analysis, especially when performing single molecule sequencing by synthesis.

Abstract: The present invention provides a method for inhibiting growth of a cancer cell, particularly a renal cell carcinoma, by contacting the cell with a composition composed of an HIG2 siRNA or HIG2 antibody. Methods of diagnosing renal cell cancer are also provided within the present invention.

Abstract: The methods of the invention detect epidermal growth factor RNA, epidermal growth factor receptor RNA, her-2/neu RNA, c-myc RNA, heterogeneous nuclear ribonucleoprotein A2/B1 RNA or any combination thereof in blood plasma, serum, and other bodily fluids. The inventive methods are useful for detection, diagnosis, monitoring, treatment, or evaluation of neoplastic disease.

Abstract: The present invention relates to a method for the diagnosis and/or the follow up of the evolution of cancer, which includes the analysis and quantification of over expressed and amplified genes in the plasma/serum of cancer patients or persons suspected to harbor cancer. This is achieved by analyzing together the amount of DNA and RNA of certain genes in the plasma/serum of cancer patients that are the reflection of a gene amplification and/or a gene over expression in comparison to healthy controls.

Abstract: The present invention provides a method of covalently joining a DNA strand to an RNA strand comprising (a) forming a topoisomerase-DNA intermediate by incubating a DNA cleavage substrate comprising a topoisomerase cleavage site with a topoisomerase specific for that site, wherein the topoisomerase-DNA intermediate has one or more 5? single-strand tails; and (b) adding to the topoisomerase-DNA intermediate an acceptor RNA strand complementary to the 5? single-strand tail under conditions permitting a ligation of the covalently bound DNA strand of the topoisomerase-DNA intermediate to the RNA acceptor strand and dissociation of the topoisomerase, thereby covalently joining the DNA strand to the RNA strand. The present invention also provides a method of tagging a 5? end of an RNA molecule. The present invention further provides a DNA-RNA molecule which has been joined in vitro by the use of a topoisomerase. The present invention also provides a method of tagging a 5? end of an mRNA.

Abstract: A method of diagnosing a disease that includes obtaining experimental data on gene selections. The gene selection functions to characterize a cancer when the expression of that gene selection is compared to the identical selection from a noncancerous cell or a different type of cancer cell. The invention also includes a method of targeting at least one product of a gene that includes administration of a therapeutic agent. The invention also includes the use of a gene selection for diagnosing a cancer.

Abstract: A method of synthesizing a cDNA chain using an insoluble carrier having on the surface thereof a polymer substance containing a first unit having a group derived from a phosphoric ester composing the hydrophilic portion of phospholipid, and a second unit having a group derived from carboxylic acid having an electron-attractive substituent bound to a carbonyl group, which includes immobilizing a polynucleotide for DNA elongation; bringing a solution containing an RNA fragment, nucleotide monomers, and a reverse transcriptase or an enzyme having polymerase activity into contact with the surface of the insoluble carrier; and allowing the polynucleotide for DNA elongation immobilized on the surface of the carrier to elongate using the RNA fragment contained in the solution as a template, to thereby form a single-strand cDNA.

Abstract: The present invention is directed generally to activating gene expression or causing over-expression of a gene by recombination methods in situ. The invention also is directed generally to methods for expressing an endogenous gene in a cell at levels higher than those normally found in the cell. In one embodiment of the invention, expression of an endogenous gene is activated or increased following integration into the cell, by non-homologous or illegitimate recombination, of a regulatory sequence that activates expression of the gene. In another embodiment, the expression of the endogenous gene may be further increased by co-integration of one or more amplifiable markers, and selecting for increased copies of the one or more amplifiable markers located on the integrated vector. In another embodiment, the invention is directed to activation of endogenous genes by nontargeted integration of specialized activation vectors, which are provided by the invention, into the genome of a host cell.

Abstract: The present invention is directed to the thermostable DNA-polymerase protein of the archaeal ampullavirus ABV (Acidianus Bottle-shaped virus) and the nucleic acid encoding said DNA polymerase. The invention also relates to method of synthesizing, amplifying or sequencing nucleic acid implementing said DNA polymerase protein and kit or apparatus comprising said DNA polymerase protein.

Abstract: Methods and kits performing reverse transcription and RT-PCR reactions having high fidelity, processivity and DNA polymerase activity are described. The methods involve performance of reverse transcription at an increased temperature with a reverse transcriptase from Simian Immunodeficiency Virus-agm.sab or a variation thereof. The kits of the present invention include a reverse transcriptase from Simian Immunodeficiency Virus-agm.sab or a variation thereof, a DNA polymerase capable of amplifying cDNA under conditions suitable for polymerase chain reaction, and the reagents necessary to carry out both processes.

Abstract: Compositions and methods are disclosed for automated storing, tracking, retrieving and analyzing biological samples, including dry storage at ambient temperatures of nucleic acids, proteins (including enzymes), and cells using a dissolvable dry storage matrix that permits recovery of biologically active materials. RFID-tagged biological sample storage devices featuring dissolvable or dissociable matrices are described for use as supports of biological samples, which matrices can be dried and subsequently rehydrated for sample recovery. Also disclosed are computer-implemented systems and methods for managing sample data.

Abstract: The present invention relates to a method for diagnosing and/or predicting a septic syndrome, wherein: a. a biological sample from the patient is made available and biological material is extracted from the biological sample b. at least four specific reagents which are selected from the following specific reagents are made available: reagent which is specific for the target gene IL-10, reagent which is specific for the target gene TGF?, reagent which is specific for the target gene HMG1, reagent which is specific for the target gene T-bet, reagent which is specific for the target gene IL-1?, reagent which is specific for the target gene TNF? and reagent which is specific for the target gene GATA-3 c. the expression of at least four target genes selected from: IL-10, TGF?, HMG1, T-bet, IL-1?, TNF? and GATA-3 is determined. The present invention also relates to a kit for diagnosing and/or predicting a septic syndrome.

Abstract: Diagnostic and therapeutic applications for certain types of cancer and precancerous conditions, including those deriving from hematologic cells, are described. Of particular interest are those cancers and precancerous conditions associated with increased signaling in the RAS-MAP kinase pathway. The diagnostic and therapeutic applications described herein are based on certain mutations in the protein tyrosine phosphatase gene PTPN11 and its expression product, PTPN11, promoting a gain-of-function in PTPN11 activity. Also described are nucleotide sequences, amino acid sequences, probes, and primers related to PTPN11 and PTPN11 variants, and cells expressing such variants.

Abstract: Double-stranded RNA (dsRNA) induces sequence-specific post-transcriptional gene silencing in many organisms by a process known as RNA interference (RNAi). Using a Drosophila in vitro system, we demonstrate that 19-23 nt short RNA fragments are the sequence-specific mediators of RNAi. The short interfering RNAs (siRNAs) are generated by an RNase III-like processing reaction from long dsRNA. Chemically synthesized siRNA duplexes with overhanging 3? ends mediate efficient target RNA cleavage in the lysate, and the cleavage site is located near the center of the region spanned by the guiding siRNA. Furthermore, we provide evidence that the direction of dsRNA processing determines whether sense or antisense target RNA can be cleaved by the produced siRNP complex.

Abstract: The present invention provides methods for providing as tags the nucleotide sequences at the 5? end of mRNA. The method of the present invention comprises the step of synthesizing cDNA using, as a template, mRNA whose CAP structure is linked with a IIs linker having a type IIs endonuclease recognition sequence. Tags including the nucleotide sequence from the 5? end of mRNA are generated by reacting the type IIs endonuclease to cDNA. Tags can be generated from all mRNA, independently of their nucleotide sequences. Methods for identifying transcriptional start sites and primers for full-length cDNA synthesis are provided based on the nucleotide sequence information of tags of the present invention.

Abstract: Cellular mRNA-protein (mRNP) complexes are partitioned in vivo by contacting a biological sample with at least one ligand that specifically binds at least one component of a mRNP complex. Suitable biological samples comprise at least one mRNA-protein (mRNP) complex and include cell cultures, cell extracts, and whole tissue, including tumor tissue. Ligands include antibodies that specifically bind RNA-binding or RNA-associated proteins present in the mRNP complex. The mRNP complex is separated by binding the ligand with a binding molecule specific for the ligand, where the binding molecule is attached to a solid support. The mRNP complex is collected by removing the mRNP complex from the solid support. After collecting the mRNP complex, the mRNA bound within the complex may be characterized and identified. Subsets of the total mRNA population of a cell may accordingly be characterized, and a gene expression profile of the cell obtained.

Abstract: The present invention relates generally to methods, compositions and kits for synthesizing sense RNA molecules from one or more RNA molecules of interest in a sample. In exemplary embodiments, the methods use a terminal tagging oligoribonucleotide (rTTO) to join a DNA sequence tag to the 3?-termini of first-strand cDNA molecules. The use of an rTTO comprising ribonucleotides results in decreased oligonucleotide-derived background synthesis of RNA in the absence of sample RNA and, surprisingly and unexpectedly, also results in significantly increased yields of sense RNA molecules that exhibit sequences that are substantially identical to those of the RNA molecules of interest in the sample. The sense RNA molecules also have an RNA sequence tag on their 5?-termini that is useful for fixing the lengths of sense RNA molecules that are synthesized in a second or subsequent round.

Abstract: This invention relates to new polypeptides which exhibit kinase activity or, more specifically, which show phosphoinositide (PI) 3-kinase activity. Such polypeptides are involved in pathways responsible for cellular growth and differentiation. An isolated polypeptide which possesses PI3-kinase activity when produced by recombinant production in insect cells is disclosed.

Abstract: The invention relates to compositions, kits, and methods for detecting, characterizing, preventing, and treating prostate cancer. FKBP markers are provided, wherein changes in the levels of expression of one or more of the FKBP markers is correlated with the presence of prostate cancer.

Abstract: This invention relates to: a labeled amino acid that can be introduced into a protein with the aid of a protein synthesis system; a functional protein having functions derived from a label compound; a labeled amino acid comprising an aromatic ring bound to an amino acid side chain and a label compound bound thereto via the aromatic ring; a functional protein to which the labeled amino acid has been introduced; and a novel method for effectively obtaining a labeled amino acid-tRNA complex.

Abstract: The present invention is directed to providing sensitive and accurate assays for gene detection, genome-wide gene expression profiling and alternative splice monitoring with a minimum or absence of target-specific amplification.

Abstract: Methods and compositions are provided for amplifying RNA from an RNA template. Methods and compositions are further provided for detecting an RNA in an RNA containing sample. Compositions used for the amplification of RNA from RNA include a RNA-dependent RNA-polymerase, a RNA helicase, and an energy source. Illustrative RNA-dependent RNA-polymerase enzymes are derived from the tomato or tobacco plant, while illustrative RNA helicase enzymes include the eIF4A and eIF4B proteins.

Abstract: This invention provides novel methods for assessing HPV infection. Gene expression levels are used to assess the progression of HPV infection from benign to malignant growth. Also provided are kits for carrying out the methods of this invention.