7 results about "Asymmetric PCR" patented technology

The present invention relates to methods and compositions for preparing a sample enriched for single stranded nucleic acids. The single stranded nucleic acids find use as templates for synthesizing Xpandomer copies of the nucleic acids. The Xpandomer copies may be passed through a nanopore sensor to determine the nucleic acid sequence of the nucleic acids. The methods include asymmetric PCR, amplicon immobilization melt, exonuclease-mediated partial or complete digestion of one or both strand of an amplicon and linear amplification.

The present invention relates to methods and compositions for preparing a sample enriched for single stranded nucleic acids. The single stranded nucleic acids find use as templates for synthesizing Xpandomer copies of the nucleic acids. The Xpandomer copies may be passed through a nanopore sensor to determine the nucleic acid sequence of the nucleic acids. The methods include asymmetric PCR, amplicon immobilization melt, exonuclease-mediated partial or complete digestion of one or both strand of an amplicon and linear amplification.

The invention discloses a universal CRISPR / Cas12a (clustered regularly interspaced short palindromic repeats / CRISPR associated 12a) system, which can realize visual rapid detection of acinetobacter baumannii drug resistance genes. The system is combined with an asymmetric PCR technology to amplify one housekeeping gene and four drug-resistant genes of bacteria to generate a single-stranded DNA product, so that the amplification efficiency and the signal intensity are remarkably improved, and a high-specificity substrate is provided for subsequent reaction. The single-chain amplicon and the modular ring engineering hairpin are subjected to a strand displacement reaction, and Cas12a protein activation is accurately regulated and controlled; the activated Cas12a cuts the reporter molecule, and the product presents a visual visual signal through a lateral flow chromatography test strip. The strategy has prominent universality, crRNA does not need to be redesigned or an activating agent does not need to be added when five targets are detected, plug-and-play can be achieved only by replacing the hairpin response sequence, and the method can be conveniently expanded to detection of other bacteria, viruses and drug-resistant genes. The system is easy and convenient to operate, high in specificity and good in stability, provides a simplified and universal CRISPR diagnosis new scheme for clinical bacterial genotyping, and has important clinical application and popularization value.

The invention discloses a PCR-CRISPR method and a PCR-CRISPR system for integrated detection of drug-resistant sites of mycobacterium tuberculosis and rifampicin. Specifically, the invention discloses a PCR-CRISPR method for in-vitro non-diagnostic and non-therapeutic integrated detection of mycobacterium tuberculosis and rifampicin drug-resistant mutation sites, and the method comprises the following steps: (s1) providing a sample to be detected, and extracting total DNA; (s2) pushing nucleic acid liquid containing the total DNA into a first reaction chamber, and carrying out asymmetric PCR reaction to obtain a first fluorescence signal; (s3) pushing a PCR amplification product in the step (s2) into a second reaction chamber, carrying out CRISPR reaction in the second reaction chamber, and obtaining a second fluorescence signal for judging whether the target sequence exists or not; and (s4) combining and analyzing the first fluorescence signal and the second fluorescence signal to obtain a detection result of the mycobacterium tuberculosis and rifampicin drug-resistant mutation site. The method disclosed by the invention is high in detection sensitivity, convenient to operate and low in cost, and can be used for effectively screening tuberculosis and multidrug-resistant tuberculosis.

The application discloses an ABO genotyping composition and kit based on a multiple asymmetric PCR probe melting curve and application. The application can realize accurate ABO genotyping in one batch of PCR reactions, and the whole detection process is simple, rapid, accurate and efficient, and has high practical value. The application makes up for the shortcomings of the existing serological detection method, has the advantages of standardization, low cost and low technical barrier for clinical popularization, and has a wide application prospect in ABO genotyping for Chinese population.

The invention provides a molecular comparator based on a miRNA pair and application of the molecular comparator. The molecular comparator comprises a miRNA marker, an asymmetric PCR reaction device, a TMSD reaction device and a fluorescence signal output device, the method comprises the following steps: sequentially passing miRNA markers through an asymmetric PCR reaction device to obtain ssDNA, placing the ssDNA in a TMSD reaction device to obtain a probe E and a probe F, hybridizing the probe E and the probe F, and carrying out threshold-based comparison on the relative abundance of miR-148a-3p and miR-30d-5p on a molecular level by using a fluorescence signal output device; the relative expression level of the specific miRNA pair is internally normalized, interference of irrelevant physiological changes is reduced, and compared with diagnosis based on the absolute content of a single miRNA, the method has more excellent performance and can be applied to preparation of a non-small cell lung cancer detection kit.

The present application relates to the application of graphene oxide in the preparation of single-stranded DNA secondary library for aptamer screening. Specifically, graphene oxide is incubated with the product after asymmetric PCR amplification, and the single-stranded DNA secondary library on graphene oxide can be obtained after centrifugation and washing. The method of the present application is simple, does not affect the specificity and affinity of aptamer, has the advantages of low cost and high efficiency, overcomes the problems of existing single-stranded DNA secondary library preparation methods, such as many steps, high operation requirements and high cost, and helps to screen high-quality aptamer.