84 results about "Oligonucleotide hybridization" patented technology

The present invention relates to a method for the comparative assessment of the level of specific nucleic acid sequences in samples derived from different sources. More specifically, the invention relates to a method using oligonucleotides covalently linked to a solid support, such as beads, to isolate specific labeled nucleic acid sequences from complex mixtures. The methods disclosed allow quantitative comparisons of the amount of nucleic acid of defined sequence in a plurality of different samples of nucleic acid, e.g., from different cells or tissues or from genetic libraries. Nucleic acids from the samples are labeled in such a fashion that the signals can be distinguished and compared following hybridization to the oligonucleotides on the beads. According to the invention, the solid supports with the hybridized nucleic acid may be retrieved, and the target nucleic acid eluted and analyzed. Furthermore, the invention provides a method for tagging individual clones from a cDNA library such that they can be identified uniquely and retrieved by hybridization to specific beads.

The present invention provides a method of detecting antigens, which comprises immobilizing an antigen to a solid support and contacting the solid support with a means for hybridizing a labeled dendrimer to the antibody, through an oligonucleotide complexed thereto. A directly oligonucleotide labeled primary antibody or an oligonucleotide labeled secondary antibody may be employed, and a conventionally labeled dendrimer can subsequently be hybridized to the oligonucleotide through one or more of the outer arms of the dendrimer. The present invention offers the advantage over conventional methods of antigen detection by providing multiple label molecules per antigen, thereby enhancing the observed signal associated with the label.

A system and method for simultaneous, quantitative measurement of nucleic acids in a sample. Fluorescently tagged amplicons of the target nucleic acids are localized on a substrate surface by hybridization to oligopobes that have been arrayed and tethered to the substrate surface in a pre-determined, two-dimensional pattern. The hybridized, amplicons are then detected by exciting their fluorescent tags using an evanescent wave of light of the appropriate wave-length. Because of the limited penetration of the evanescent wave (about 100-300 nm), the fluorescently tagged nucleotides in the remainder of the reaction cell do not fluoresce. By measuring the fluorescence at various locations on the substrate surface, the current abundance of hybridized amplicons of each of the target nucleic acids can be determined. The analytic techniques of real time PCR may then be used to obtain accurate, quantitative measurements for each of the nucleic acids in the sample.

Disclosed are methods for detecting a target nucleic acid molecule having a known sequence among a plurality of nucleic acid molecules. The method includes hybridizing two oligonucleotides to adjacent nucleotide sequences on the target nucleic acid molecule, ligating the two oligonucleotides together, and extending a primer that hybrizes to the 3′ end of the oligonucleotide that hybrized to the 5′ nucleotide sequence on the target nucleic acid molecule. If the target nucleic acid molecule is present (and thereby allowing ligation of the two oligonucleotides), extension of the primer dislodges a detection probe hybridized to the 5′ end of the oligonucleotide that hybridized to the 3′ nucleotide sequence on the target nucleic acid molecule.Also disclosed are kits for performing the methods of the invention.

The present invention relates to a method for detecting an analyte in a sample, said method comprising: (a) contacting said sample with at least one set of at least first and second proximity probes, wherein said probes each comprise an analyte-binding moiety and can simultaneously bind to the analyte, and wherein (i) said first proximity probe comprises a nucleic acid moiety attached at one end to the analyte-binding moiety, wherein a circular or circularizable oligonucleotide is hybridized to said nucleic acid moiety before, during or after said contacting step; and (ii) said second proximity probe comprises an enzyme moiety, attached to the analyte-binding moiety, capable of directly or indirectly enabling rolling circle amplification (RCA) of the circular or, when it is circularized, of the circularizable oligonucleotide hybridized to the nucleic acid moiety of the first proximity probe, wherein said RCA is primed by said nucleic acid moiety of said first proximity probe; (b) if necessary, circularizing said oligonucleotide, to produce a circularized template for RCA; (c) subjecting said circular or circularized template to RCA, wherein if the enzyme moiety of the second proximity probe in step (a)(ii) is a DNA polymerase, this step does not utilize a free DNA polymerase; and (d) detecting a product of said RCA.

The present invention relates to the detection of a target nucleic acid sequence by a PTOCE (PTO Cleavage and Extension) assay. The present invention detects a target nucleic acid sequence in which the PTO (Probing and Tagging Oligonucleotide) hybridized with the target nucleic acid sequence is cleaved to release a fragment and the fragment is hybridized with the CTO (Capturing and Templating Oligonucleotide) to form an extended duplex, followed by detecting the presence of the extended duplex. The extended duplex provides signals (generation, increase, extinguishment or decrease of signals) from labels indicating the presence of the extended duplex and has adjustable Tm value, which are well adoptable for detection of the presence of the target nucleic acid sequence.

It is an object to provide a method of suitably analyzing the amount of gene expression of a single-cell.A method of detecting a nucleic acid comprisinga step of sampling a single-cell from a sample containing at least a single-cell,a cell lysis step of lysing cell membrane of the sampled single-cell and extracting nucleic acids from the cell,a DNase treatment step of degrading DNA of the extracted nucleic acids with DNase,a step of hybridizing mRNA of the total RNA contained in the single-cell with oligo (dT) fixed onto a carrier,a step of performing reverse transcription of the mRNA hybridized with the oligo (dT) to fix cDNA derived from the single-cell onto the carrier, thereby preparing a single-cell derived cDNA library fixed onto a carrier, anda step of amplifying cDNA fixed onto the carrier and simultaneously detecting an amplification amount of the cDNA.

The invention provides a novel method of labeling oligonucleotides, with reporter moieties, including but not limited to, quenchers, fluorophores, biotin, digoxigenin, peptides and proteins. In addition, this invention provides a method of detecting hybridization of oligonucleotides. This invention also provides novel azo quenchers having the general formula shown below.The invention further provides compositions comprising labeled oligonucleotides and solid supports. The invention also provides kits comprising at least one composition of the present invention.

The present invention relates to prostate cancer. More specifically, the present invention relates to a method to detect prostate cancer in a patient sample by detecting the RNA encoded by the gene PCA3. More particularly the present invention relates to a method for determining a predisposition, or presence of prostate cancer in a patient comprising: (a) contacting a biological sample of a patient with at least one oligonucleotide that hybridizes to a PCA3 polynucleotide; (b) detecting in the biological sample an amount of PCA3 and second prostate specific polynucleotides; and (c) comparing the amount of PCA3 polynucleotide that hybridizes to the oligonucleotide to a predetermined cut off value, and therefrom determining the presence or absence of prostate cancer in the biological sample. The present invention further relates to diagnostic kits for the detection of prostate cancer or the risk of developing same in a patient comprising: (a) at least one container means having disposed therein at least one oligonucleotide probe or primer that hybridizes to one a PCA3 nucleic acid or complement thereof; (b) at least one oligonucleotide probe or primer that hybridizes with a second prostate specific nucleic acid or complement thereof; and (c) reagents enabling a detection of PCA3 and of the second prostate specific nucleic acid when PCA3 or second prostate-specific nucleic acid sequence is present.

Solid support assays using non-standard bases are described. A capture oligonucleotide comprising a molecular recognition sequence is attached to a solid support and hybridized with a target oligonucleotide. In some instances, the molecular recognition sequence includes one or more non-standard bases and hybridizes to a complementary tagging sequence of the target oligonucleotide. In other instances, incorporation of a non-standard base (e.g., via PCR or ligation) is used in the assay.

The invention provides assays that can detect multiple genetic variants of a gene (e.g., a mycobacterial gene) in a sample using a pool (e.g., 2, 3, 4, or more) of oligonucleotide hybridization probes.

A method for measuring a test substance whereby the test substance in a sample can be specifically measured without using an antibody against the test substance; an aptamer molecule used therefor; and a method for creating the aptamer are disclosed. An aptamer capable of hybridizing with an oligonucleotide when it is bound to a test substance, but is incapable of hybridizing with the oligonucleotide when it is not bound to the test substance, is utilized. The aptamer is brought into contact with a sample, and the aptamer bound to the test substance is brought into contact with an immobilized oligonucleotide which hybridizes with the aptamer, to bind the aptamer to a solid phase, followed by measurement of the aptamer immobilized on the solid phase.

The present invention relates to the detection of a target nucleic acid sequence by a PCE-SH (PTO Cleavage and Extension-Dependent Signaling Oligonucleotide Hybridization) assay. The present invention does not use probes to be hybridized with target nucleic acid sequences for providing target signals. Interestingly, the present invention uses probes (signaling oligonucleotides) to be hybridized with the extended strand formed in a target-dependent manner in which the extended strand is synthesized using the CTO artificially selected as templates.

The invention provides a method for carrying out allele variant specific detection by utilizing an inhibitor, a kit and a composition. The method provided by the invention comprises the following steps of: adding a primer pair and an inhibitor the terminal 3' of which can not be extended by polymerase in a hybridization process into a nucleic acid sample, wherein the primer pair comprises a specific primer and a reverse primer; hybridizing the specific primer and the reverse primer respectively with target sequence mutant type oligonucleotide in the nucleic acid sample to produce an amplicon; hybridizing the inhibitor with a target sequence wild type oligonucleotide in the nucleic acid sample; and carrying out real-time specific detection on a mutant type allele by utilizing a fluorescent dye or a detection pointer. The added inhibitor can not extend in an amplification process, and amplification of a wild type allele variant is inhibited, so that the amplification efficiency of a mutant type allele variant is improved, and the sensitivity and specificity of detection are improved.

A disposable sample processing module is provided for processing DNA or RNA samples. The module includes a hybridization chamber adapted to receive an oligonucleotide covalently bonded to an internal surface of the hybridization chamber. The module also include a sample well adapted to hold a DNA or RNA sample, said sample well being coupled to the hybridization chamber, a moveable valve plate disposed between the sample well and hybridization chamber, said moveable valve plate having a first position that allows transfer of the DNA or RNA sample from the sample well to the hybridization chamber and a second position that blocks transfer to the hybridization chamber and a manifold adapted to exchange fluids with the hybridization chamber to hybridize the DNA or RNA sample with the oligonucleotide, to wash the hybridized sample and to amplify the hybridized sample.

The present invention is in the field of chemistry and biotechnology. The present invention relates to cell-based microarrays, improved methods for forming such arrays, and methods for using such arrays in diagnostics, therapeutics and research. The invention particularly concerns microarrays in which ligands of a target cells are immobilized to the array support via ligand-binding molecules bound to an oligonucleotide that is hybridized to a support-immobilized oligonucleotide.

The present invention relates to the detection of a target nucleic acid sequence by a PTOCE (PTO Cleavage and Extension) assay. The present invention detects a target nucleic acid sequence in which the PTO (Probing and Tagging Oligonucleotide) hybridized with the target nucleic acid sequence is cleaved to release a fragment and the fragment is hybridized with the CTO (Capturing and Templating Oligonucleotide) to form an extended duplex, followed by detecting the presence of the extended duplex. The extended duplex provides signals (generation, increase, extinguishment or decrease of signals) from labels indicating the presence of the extended duplex and has adjustable Tm value, which are well adoptable for detection of the presence of the target nucleic acid sequence.

Compositions and methods for isolating new variants of known gene sequences are provided. The methods find use in identifying variants, particularly homologs, in complex mixtures. Compositions comprise hybridization baits that hybridize to gene families of interest, particularly agricultural interest, in order to selectively enrich the polynucleotides of interest from complex mixtures. Bait sequences may be specific for a number of genes from distinct gene families of interest and may be designed to cover each gene of interest by at least 2-fold. Thus methods disclosed herein are drawn to an oligonucleotide hybridization gene capture approach for identification of new genes of interest from environmental samples. This approach bypasses the need for labor-intensive microbial strain isolation, permits simultaneous discovery of genes from multiple gene families of interest, and increases the potential to discover genes from low-abundance and unculturable organisms present in complex mixtures of environmental microbes.

An immobilized bimolecular construct comprises a solid support, a first oligonucleotide and a second oligonucleotide. The first oligonucleotide is labeled at one end with a fluorophore or quencher and attached at the other end to a solid support. The second oligonucleotide is labeled at one end with a fluorophore or quencher and hybridized at the other end to the first oligonucleotide. Hybridization of the second oligonucleotide with the first oligonucleotide brings the labeled end of the second oligonucleotide in close proximity or physical contact with the labeled end of the first oligonucleotide. In one embodiment the second oligonucleotide is also attached to the solid support in proximity to the first oligonucleotide. In this embodiment, the second oligonucleotide may be first attached to the solid support and then hybridized to the first oligonucleotide or, conversely, first hybridized to the first oligonucleotide and then attached to the solid support.

The invention relates to an oligonucleotide microarray technique for detecting pathogen contamination in seawater, belonging to the field of seawater contamination monitoring. The technique comprises the main technical schemes that a 16S-23S rRNA gene transcription interval sequence is used as a detection target and is amplified by a one-step polymerase chain reaction, a digoxin mark is obtained simultaneously, and then oligonucleotide hybridization is carried out; and the obtained monitoring result is interpreted in a manner that an enzyme-labeled antibody catalyzes the substrate colour development. Compared with the traditional product for detecting seawater contamination, the invention utilizes microarray detection to obtain the distribution situation of large numbers of pathogens and contamination index bacteria, and has the advantage of high flux; the invention can directly utilize seawater as a sample and truly obtain the contamination situation information of target bacteria under a condition of keeping the natural proportion of the flora number in the seawater; however, most existing detection techniques need the step of enrichment culture, destroy the original proportion of a flora composition, have lower reliability of the result and have longer detection procedure; and the oligonucleotide microarray detection operation has short procedure and is comparatively sensitive and fast.

The present invention relates to methods for the extraction of long fragment RNA from fixed tissue specimens. In particular, the present invention relates to methods for the extraction of RNA from formalin-fixed paraffin-embedded tissue specimens for use in biologic applications, including assays based on oligonucleotide hybridization.

The present invention provides a method for analysing the interaction of one or more nucleotide sequence(s) from one or more region(s) of interest with other nucleotides sequences in a three-dimensional DNA structure, comprising the steps of: (a) providing a sample of crosslinked DNA; (b) digesting the cross-linked DNA with a first restriction enzyme; (c) ligating the cross-linked nucleotide sequences; (d) reversing the cross-linking; e) fragmenting the ligation and ligated molecules from (d); (f) hybridising the fragments from (e) to one or more oligonucleotides representing the sequences which are adjacent to the cleavage site of the first restriction enzyme in order to enrich for the ends of the nucleotide sequences that have been ligated to another nucleotide sequence in step (c); and (g) analysing the nucleotide sequence of the enriched fragments in order to identify the nucleotide sequences involved in interaction(s).

The invention discloses a detection method synthesizing microfluidic chips in situ, which comprises the following steps that: 1) a sequence of non-coding small RNAs reported on the prior model plant is utilized, sequence information of miRNA is preferably selected, and complementary oligonucleotide hybridization probes are designed according to the sequence of a sense strand or an antisense strand of miRNA; 2) an oligonucleotide chip is prepared by an in-site synthetic method; 3) small RNAs smaller than a 50 oligonucleotide group are extracted from a plant to be detected, then are hybridized with the probes in the chip after making end labeling, and the result of chip hybridization is obtained to perform qualitative and quantitative analysis. The detection method can be used for the validation of miRNA (and other sNC-RNAs) obtained through the forecast by a bioinformatics method, can also be used for the detection and identification of miRNA (and other sNC-RNAs) in new plant species, and can also be used for the determination of miRNA (and other sNC-RNAs) variation relation under different treatment conditions (eg. under the condition of virus infection).