21 results about "Molecular beacon" patented technology

The present application relates to the field of molecular biology, in particular to a primer probe composition, kit and detection method for 23 human papilloma virus genotyping detection, wherein the hydrolysis probe and molecular beacon probe are respectively designed in two different regions of HPV gene sequence E6 and L1, and the fluorescence collection point in the amplification stage is set in the high-temperature denaturation stage, so that the detection in the amplification stage and the melting curve stage is independent and does not interfere with each other, thereby realizing the combined use of real-time fluorescent PCR amplification detection technology and melting curve technology; the technology improves the single-well detection target flux of fluorescent PCR analysis technology, reduces the detection cost, and provides a more optimal detection method for HPV multiplex fluorescent PCR genotyping detection.

The invention relates to the technical field of biology, and discloses a molecular beacon and a high-throughput sequencing library absolute quantification method thereof, and a fluorescent molecular beacon oligonucleotide (Oligo) sequence comprises a nucleotide chain (Loop chain) of a sequencing library recognition region, a first universal sequence region at the 5'upstream of the nucleotide chain (Loop chain) of the sequencing library recognition region, and a second universal sequence region at the 3 'downstream of the nucleotide chain (Loop chain) of the sequencing library recognition region; 5'of the first universal sequence region is modified into a fluorophore, and 3 'of the second universal sequence region is modified into a quenching group; and a nucleotide chain of the sequencing library recognition region is from an Illumina anmena sequencing platform or an MGI Huazai sequencing platform. The invention comprises a fluorescent molecular beacon for high-throughput sequencing library sequencing before-loading quantification, a sequence applied to a high-throughput sequencing platform and a detection method for library absolute quantification based on the molecular beacon, and overcomes the defects of a current gold standard detection method in technology and efficiency. The quantitative precision of the sequencing library is ensured; meanwhile, the economic and time cost of an actual application end is greatly reduced.

The application provides a probe combination, a kit and a detection method for detecting nucleic acid, and belongs to the technical field of nucleic acid detection. The application provides a probe combination for detecting nucleic acid, wherein the probe combination comprises a specific hairpin switch probe HSP and a universal molecular beacon; the HSP is composed of a middle template binding sequence, two end beacon binding sequences and complementary sequences. The universal molecular beacon comprises a short stem structure and a long loop sequence, and the loop sequence is designed to be complementary to the beacon binding sequence region of the HSP. When the temperature is increased or there is a DNA strand complementary to the middle loop sequence, the universal molecular beacon is unfolded from a hairpin structure to a linear structure, the 5' and 3' end groups are far away from each other, and the fluorescence is significantly increased. The HSP and the universal molecular beacon provided by the application are suitable for the detection of qPCR and ddPCR, and can meet the detection of nucleic acid and mutant nucleic acid, and have high sensitivity and strong specificity.

The invention discloses a universal molecular beacon kit for various animal-derived components, and belongs to the technical field of molecular biological detection. Aiming at gene segments with specific structural characteristics in 11 common livestock and poultry animal genomes, the invention designs a composition of a universal primer and a molecular beacon fluorescent probe. The combination can be used for carrying out real-time fluorescent PCR detection and melting curve analysis by using the kit and the method by taking genome DNA of a sample to be detected as a template. By comparing the shape of a melting peak of a detected sample and the characteristic melting temperature (Tm value) of the detected sample, 11 animal-derived components of livestock and poultry can be quickly and accurately identified under the condition of single-tube detection. Technical innovation is achieved in the field of molecular biological detection, an efficient and reliable solution is provided for animal-derived component detection, and the method has important practical value and popularization potential.

The application belongs to the technical field of biomedicine, and relates to a mitochondrion-targeted nano device and a preparation method and application thereof, the nano device comprising a cascade-activated photodynamic molecular beacon and a mitochondrion-targeted delivery carrier, the cascade-activated photodynamic molecular beacon comprising an APE1 cleavage site and a microRNA recognition region. The cascade-activated photodynamic molecular beacon and the mitochondrion-targeted delivery carrier are self-assembled into a complex through electrostatic adsorption, and cascade activation and signal amplification are realized through the dual response of miR-146a / APE1 which is highly expressed in mitochondria of tumor cells. The application triggers precise and controllable cell pyroptosis through tumor-specific signals, reverses the immune-inhibitory microenvironment, converts the 'cold tumor' into a 'hot tumor', significantly improves the effect of cancer immunotherapy, has good biocompatibility and tumor selectivity, and provides a new strategy and tool for precise treatment of cancer.

The application discloses a kind of digital single molecule enzyme multi-activity characteristic cross-scale heterogeneity dynamic evaluation methods, comprising: the enzyme-template-primer ternary complex of pre-combination completion is diluted to single molecule level to be loaded into the reaction chamber in microwell array chip to carry out rolling circle amplification, and utilize molecular beacon and the fluorescence signal released by amplification product combination;Real-time monitoring and obtaining the fluorescence signal to generate time-resolved fluorescence intensity curve;The slope parameter of the fluorescence intensity curve is extracted to obtain the catalytic synthesis rate by conversion calculation for evaluating enzyme activity;The catalytic synthesis rate value distribution is fitted using n times Gaussian mixture model to obtain the standard deviation for evaluating population uniformity and the coefficient of variation for evaluating stability.The application can realize single molecule resolution, multi-parameter dynamic correlation and environmental interference processing cross-scale analysis simultaneously, and break through the resolution limit of traditional analysis technology by integrating single molecule fluorescence tracking and population statistics.

The present invention provides novel primers and sloppy molecular beacons and molecular beacon probes for amplifying segments of different genes in MycobacteriumTuberculosis to identify the presence of M. tuberculosis DNA and / or to identify resistance to anti-tuberculosis drugs.SOLUTION: Oligonucleotide sets are provided for amplifying a portion of an M. tuberculosis region selected from the group consisting of rpoB, gyrA, gyrB, inhA promoter, rrs, eis promoter, embB, katG, dosR, IS6110, and IS1081 genes.SELECTED DRAWING: None

The present application is related to a method for in situ detection of multiple lung cancer-related markers in live circulating malignant cells, and tumor heterogeneity, including accurately detecting at least one lung cancer-related nucleic acid marker in live lung cancer cells by using a lung cancer-targeted nanoprobe loaded with a plurality of molecular beacons to reflect levels of different markers in the lung cancer cells from a single cell level and a heterogeneous state of a tumor of a specific patient. The nanoprobe is a nanoparticle self-assembled from a high polymer material, an electropositive protein, a functional polypeptide and / or a functional aptamer, and a molecular beacon of a lung cancer-related marker. The nanoprobe can be used to target different phenotype circulating malignant cells in whole blood, detect nucleic acid markers in living cells, and reflect levels of different markers in the malignant cells from a single cell level.

The invention belongs to the technical field of nucleic acid molecule detection, and particularly relates to an enzyme-free isothermal strand displacement cascade amplification reagent for acute respiratory pathogen detection and application of the enzyme-free isothermal strand displacement cascade amplification reagent. The detection method based on the amplification reagent is based on a nucleic acid reaction system composed of an initiation double strand and two molecular beacons, cyclic amplification is performed through a toehold-mediated strand displacement reaction trigger signal under a constant temperature condition, and end point interpretation is performed through a fluorescence signal. The method does not need enzyme and thermal cycle, is short in detection time, adapts to high-throughput operation of a porous plate, and realizes specific recognition and signal cascade amplification of target nucleic acid under a constant temperature condition, so that rapid, accurate and high-throughput detection of acute respiratory pathogens is realized on the premise of not depending on enzymatic reaction and thermal cycle equipment. In addition, the kit has the advantages of low cost, good stability and wide application range, and is suitable for rapid detection of various acute respiratory pathogens.

The invention discloses an aptamer-mediated EXPARs-molecular beacon fluorescent biosensor as well as a preparation method and application of the aptamer-mediated EXPARs-molecular beacon fluorescent biosensor. The construction and detection process of the sensor comprises three key steps: protein binding aptamer, exponential isothermal amplification (EXPAR) and molecular beacon fluorescence resonance energy transfer (FRET). Through the ingenious design of the EXPAR system, the system realizes triple signal amplification, the detection sensitivity is remarkably improved, and Abeta42 and Abeta40 amyloid proteins as low as 100fM level can be detected. Besides, based on the sequence specificity of the aptamer and the highly specific design of each nucleic acid sequence involved in the detection system, the method can be used for simultaneously detecting the Abeta42 amyloid protein and the Abeta40 amyloid protein or other substances in the same system (such as a plasma sample), so that the detection efficiency is improved.

This invention discloses a method and kit for quantitative detection of circularized rolling circle amplified miRNAs. The method utilizes T4 RNA ligase to ligate the 3'-OH and 5'-phosphate ends of the target miRNA to form a circular miRNA. Using the circularized miRNA as a template, isothermal rolling circle amplification is performed using phi29 DNA polymerase to generate a large number of single-stranded DNA sequences with repetitive sequences. A universal molecular beacon is used to monitor the fluorescence signal of the amplification products in real time, achieving highly sensitive quantitative detection of miRNAs. This method has advantages such as high sensitivity, high specificity, and a wide linear detection range. It eliminates the need to design specific reverse transcription primers for each miRNA, can be extended to multiplex detection, and is suitable for clinical applications such as early tumor diagnosis, drug efficacy evaluation, and single-cell miRNA expression profiling.

The invention discloses an addressable nanoliter reactor chip and application of a nucleic acid signal amplification integrated system in single cell level detection markers. The nucleic acid signal amplification integrated system comprises aptamer DNA molecules and probe DNA molecules of a hairpin structure. According to the invention, the programmable characteristic of nucleic acid molecules is utilized to construct an aptamer-probe DNA synergistic signal amplification system; through combination of molecular beacon design and endonuclease reaction, in-situ signal multiplication of EpCAM protein is realized on a single cell level. After the aptamer recognizes the target epitope, the cascade hybridization reaction of the probe DNA is triggered, so that a single antigen binding event is converted into hundreds of fluorescent signal molecules to be released, and the signal gain is remarkably improved compared with that of a traditional immunofluorescence method.

The invention relates to the technical field of biosensors, in particular to a transgenic corn CP4-EPSPS protein aptamer molecular beacon fluorescent biosensor. According to the invention, a systematic aptamer rational cutting and conformation adaptation strategy is constructed, the strategy obtains an initial aptamer through magnetic bead-SELEX screening, and the CP4-EPSPS protein aptamer is precisely cut and optimized by adopting the steps of loop region active core positioning, stem region structure optimization and base binding domain identification, so that the binding performance and the structural stability are remarkably improved. Subsequently, on the basis of stable conformational characteristics, a complementary chain competition sensing mechanism is designed, and a transgenic corn CP4-EPSPS protein aptamer molecular beacon fluorescent biosensor is successfully constructed, so that amplification-free high-sensitivity rapid detection of CP4-EPSPS protein is realized, and a solution is provided for supervision of transgenic crops.

The invention discloses a technology for detecting target nucleic acid and gene mutation by using a Taq DNA polymerase large fragment (Taq KF enzyme) without 5'exonuclease activity, heat-resistant ribonuclease HII (RNase HII), and a Taqman probe or a molecular beacon with an RNA single basic group in the middle. Comprising the following steps: step 1, preparing a Taqman probe or a molecular beacon with an RNA basic group in the middle; 2, establishing a PCR amplification reaction system; 3, carrying out a PCR conventional amplification reaction and a fluorescence signal amplification reaction of target nucleic acid; 4, quantifying the concentration of the target nucleic acid in real time; and 5, determining a gene mutation type. The method has the beneficial effects that compared with the existing nucleic acid detection and gene mutation typing technology based on the conventional Taqman probe or molecular beacon, the method has obvious progress in the aspects of accuracy, sensitivity and the like.

This invention belongs to the medical field and provides a calcium carbonate manganese nanoprobe for in situ imaging detection of vascular epidermal growth factor, its preparation method, and its application. In this strategy, a DNA strand (P1 and P2) containing a protein recognition antibody, a protein recognition sequence, and a deoxyribonuclease, along with a molecular beacon DNA strand (H1), is stirred in a DMEM cell culture medium containing calcium chloride and manganese chloride to prepare the calcium carbonate manganese nanoprobe. In acidic tumor cells, this nanoprobe releases P1, P2, H1, and manganese ions. P1 and P2 can simultaneously recognize proteins, forming a complete deoxyribonuclease center, which catalyzes the cleavage of H1 under the action of the cofactor manganese ions, generating a fluorescent signal. This strategy achieves specific in situ dynamic imaging of low-abundance proteins in vivo and has great potential in biomedical research.

The invention discloses a deafness gene detection kit based on a molecular beacon and a medium probe and application, and relates to the technical field of molecular biology. The kit is used for single-tube detection of 15 mutation sites on four hereditary deafness genes, the detection kit comprises freeze-dried microspheres serving as standardized solid-state reagent units, the freeze-dried microspheres comprise specific primers, specific molecular beacons and specific medium probes, nucleotide sequences of the specific primers are shown as SEQ ID NO.35-SEQ ID NO.50, and nucleotide sequences of the specific molecular beacons and the specific medium probes are shown as SEQ ID NO.35-SEQ ID NO.50. The nucleotide sequences of the specific molecular beacons are as shown in SEQ ID NO.1 to SEQ ID NO.4, and the nucleotide sequences of the specific medium probes are as shown in SEQ ID NO.5 to SEQ ID NO.34. Compared with a traditional PCR technology, a chip technology and the like, the detection method provided by the invention can realize single-tube detection of different deafness genes, is simpler to operate, short in detection period and low in cost, and remarkably improves the detection sensitivity and accuracy.

This application discloses a technique for detecting target nucleic acid molecules using an activated fluorescent RNA probe, its expression vector, and a detection method. It includes: probe design and preparation methods; labeling and detection of target nucleic acid molecules in solution samples using the probe; labeling and detection of target nucleic acid molecules in living cells using the probe; and labeling and detection of target nucleic acid molecules in immobilized cells using the probe. This invention overcomes the shortcomings of current commercially synthesized FISH and molecular beacon probes, such as their inability to be genetically encoded, difficult preparation, and high cost. It provides an efficient, simple, and rapid method to obtain activated fluorescent RNA probes for detecting target nucleic acid molecules through genetic encoding or in vitro transcription. This offers a practical tool for research on fundamental life science issues and for point-of-care and even real-time diagnosis of diseases, possessing broad scientific and social significance.

The invention provides a high-fidelity Cas9 variant, a tissue targeting lipid nanoparticle delivery system, a CRISPR-Cas9-mediated genome precise editing method and application thereof, and belongs to the technical field of genetic engineering, the high-fidelity Cas9 variant HyperFi-Cas9 is adopted, four mutations of R691A, N497A, R661A and Q695A are introduced, and the off-target effect is reduced to 0.025% or below while the wild type targeting activity is kept not lower than 85%; a tissue targeting intelligent lipid nanoparticle delivery system is adopted, and by optimizing lipid composition and ligand modification, 30-60% of cell delivery efficiency and 16-80% of in-vivo editing efficiency are achieved; the integrated double-fluorescence real-time monitoring system comprises an FRET (fluorescence resonance energy transfer) molecular beacon probe and an mCherry-P2A-EGFP (enhanced green fluorescent protein) integrated report module, and is used for realizing dynamic monitoring and quantitative evaluation of an editing process; and a homologous recombination enhancement strategy is adopted.

The invention belongs to the technical field of biological detection, and particularly relates to a molecular beacon probe composition, a reagent and a kit for detecting miR-122. The composition comprises a molecular beacon and a specific hairpin probe, the nucleotide sequence of the molecular beacon is shown as SEQ ID NO.1, and the molecular beacon is modified with a fluorescence reporter group and a quenching group; the hairpin probe comprises two restriction enzyme cutting sites, two segments of repetitive sequences and a recognition site capable of being specifically combined with miR-122. According to the application, miR-122 is specifically combined with a hairpin probe to start a reaction, a dual-cycle cascade mechanism of target triggering, initiation chain generation and secondary chain displacement is constructed, and rapid quantitative analysis of miR-122 is realized in a single tube by a one-step method through high-fidelity recognition and exponential signal amplification of an initiation chain and a molecular beacon. The system has ultrahigh sensitivity, single-base resolution specificity and high accuracy, provides key technical support for noninvasive early screening of diseases such as liver cancer and the like, and has important clinical application value.

The invention provides a primer and molecular beacon probe combination as well as application, a kit and a detection method thereof, and relates to the technical field of biology. The primer and molecular beacon probe combination provided by the invention is high in sensitivity and strong in specificity, can be used for detecting quinolone antibiotic resistance genes pmrA, qnrS and oqxA, is more accurate and reliable in detection result, and overcomes the defects of low detection sensitivity and poor repeatability of the detection result of a traditional reagent.

The invention discloses a universal pathogen detection primer pair, a molecular beacon design method, an amplification method and application. The method comprises the following steps: synthesizing a to-be-detected pathogen universal sequence, and designing a universal upstream primer and a universal probe according to the universal sequence; the method comprises the following steps: designing a semi-specific downstream primer according to a target sequence of a to-be-detected pathogen and a universal sequence of the to-be-detected pathogen, on the basis of keeping a universal probe sequence unchanged, adding a 4-6bp sequence which is reversely complementary with a 3'end at a 5 'end of a universal probe to obtain a molecular beacon, and in PCR amplification, adding the universal sequence into the target sequence by adopting T4 DNA polymerase, by adopting a universal primer and probe design method, the workload of redesigning and verifying the primer and the probe aiming at each sample to be detected can be reduced, so that a primer and probe design and verification system aiming at different projects becomes simple, the research and development system of the projects is universalized, and the research and development efficiency is improved. The research and development process is simplified; the research and development period is shortened; and the condition optimization process is simplified.