43 results about "DNA glycosylase" patented technology

The present invention is drawn to a method for characterising nucleic acid molecules, which comprises the steps of: i) introducing a modified base which is a substrate for a DNA glycosylase into a DNA molecule; ii) excising the modified base with the DNA glycosylase to generate an abasic site; iii) cleaving the DNA at the abasic site to generate and release an extendible upstream DNA fragment having a 3′ hydroxyl terminus; and iv) incubating the released extendible upstream DNA fragment in the presence of an enzyme allowing for extension thereof and an additional template nucleic acid and analysing resultant fragment(s).

The present invention provides a method of modifying a targeted site of a double stranded DNA, including a step of contacting a complex wherein a nucleic acid sequence-recognizing module that specifically binds to a target nucleotide sequence in a selected double stranded DNA and DNA glycosylase with sufficiently low reactivity with a DNA having an unrelaxed double helix structure (unrelaxed DNA) are bonded, with the double stranded DNA, to convert one or more nucleotides in the targeted site to other one or more nucleotides or delete one or more nucleotides, or insert one or more nucleotides into the targeted site, without cleaving at least one strand of the double stranded DNA in the targeted site.

The present disclosure encompasses systems, and their methods of use, for detecting a target analyte, the systems comprising: (a) a first oligonucleotide probe comprising a reporter oligonucleotide-binding arm complementary to the nucleotide sequence of a first region of a reporter oligonucleotide and an analyte-binding arm selectively binding to a first region of a target analyte, a second oligonucleotide probe comprising a reporter oligonucleotide-binding arm having a nucleotide sequence complementary to the nucleotide sequence of a second region of a reporter oligonucleotide, and an analyte-binding arm having a nucleotide sequence characterized as selectively binding to a second region of a target analyte; and a reporter oligonucleotide comprising a region complementary to the reporter oligonucleotide-binding arm of the first oligonucleotide probe, a second region complementary to the reporter oligonucleotide-binding arm of the second oligonucleotide probe, a fluorophore and a quencher disposed on the reporter oligonucleotide and a cleavable site disposed between the fluorescent label and the quencher. The cleavable site of the reporter oligonucleotide when in the presence of a target analyte can be cleaved by an enzyme such as a restriction endonuclease, an RNase H, a Flap-endonuclease-1 (FEN-1), and a DNA glycosylase.

The invention provides a fluorescent polymerase chain reaction (PCR) kit for detecting CYP2C19 genotypes, and belongs to the field of in-vitro nucleic acid testing. The fluorescent PCR kit comprises PCR reaction liquids for detecting CYP2C19*2 and CYP2C19*3 genotypes, Taq DNA polymerase, and uracil-DNA glycosylase, wherein the PCR reaction liquids for detecting the CYP2C19*2 and CYP2C19*3 genotypes respectively comprise PCR amplification primers, minor groove binder (MGB) probes and the like; and nucleotide sequences for detecting the CYP2C19*2 and CYP2C19*3 genotypes are shown as SEQ ID NO:3-4 and SEQ ID NO:5-6 respectively. The kit has high sensitivity and specificity, can monitor the reaction progress in real time, ensures short reaction time, avoids subsequent treatment, can avoid reaction product pollution to the greatest extent, and can replace the traditional protein detection or the common PCR detection to diagnose the CYP2C19 genotypes.

The invention provides a sequencing primer for qualitative detection of cytochrome oxidase CYP2C19 genetic typing and a kit of the sequencing primer and belongs to the field of external nucleic acid detection. The kit comprises uracil DNA (Deoxyribonucleic Acid) glycosylase, Taq polymerase, PCR (Polymerase Chain Reaction) reaction liquor, PCR amplification primers, pyrophosphoric acid sequencing primers and positive control products. The kit is high in sensitivity and good in specificity, PCR products can be simply treated for a pyrophosphoric acid sequenator for sequencing, operation is simple, reaction time is short, the sensitivity of the kit is higher than that of a golden standard-capillary electrophoresis sequencing, and the kit is suitable for use for mutation analysis.

The invention relates to a sequencing primer pair for detecting an ALDH2 (Aldehyde Dehydrogenase 2) genotype with a pyrosequencing method and a kit thereof, and belongs to the technical field of in-vitro nucleic acid detection. The primer pair comprises a positive amplification primer, a reverse amplification primer and a sequencing primer, wherein a 5' end of the positive amplifier primer is subjected to biotin labeling. The kit comprises the positive amplification primer, PCR (Polymerase Chain Reaction) reaction liquid containing the reverse amplification primer, the sequencing primer, uracil DNA (Deoxyribose Nucleic Acid) glycosylase and Taq polymerase. The kit provided by the invention has the advantages of accurate detection result, high specificity, short detection period, easiness in operation, and capability of effectively meeting clinical examination requirements. Moreover, the kit further has the advantages that a reaction process can be monitored in real time, the reaction time is short, a PCR product can be subjected to pyrosequencing by a pyrosequencing instrument and high-flux sample detection through simple treatment, and the sensitivity is higher compared with a golden standard method, namely, a capillary electrophoresis sequencing method.

The invention discloses a method for ultra-sensitively simultaneously detecting multiple DNA glycosylases by using intrinsic fluorescent nucleotide. The method is a fast and sensitive fluorescence method for simultaneously detecting multiple DNA glycosylases by adopting 2-aminopurine and pyrrole-deoxycytidine as fluorophores and a DNA molecule as an intrinsic quenching agent and combining excision enzyme-assisted circulating signal amplification; and extra fluorophore and quenching group are not needed for marking, so that the target of simple, intuitive and high-sensitivity detection of an actual sample is achieved, and above all, simultaneous detection of multiple DNA glycosylases is achieved.

The invention belongs to the field of biotechnology, and particularly relates to a method for qualitatively detecting HLA-B*1502 gene with a PCR-SSP method and a clinical detection kit. The method comprises the following steps of: finding 6 specific areas capable of effectively identifying HLA-B*1502 allele type; designing 6 specific primers covering the 6 specific areas, and screening out a high-specificity primer pair applicable to PCR-SSP; and adding dUTP and UDG enzyme (uracil-DNA glycosylase) into a reaction system to solve the problem of PCR cross pollution and further improve the detection reliability. An HLA-B*1502 quick and convenient qualitative detection kit is researched and developed accordingly. The method and the clinical detection kit provided by the invention have the advantages of convenience in operation, short time, strong specificity, high accuracy, low cost and the like, and is suitable for realizing individual safe and reasonable drug use through HLA-B*1502 genotype detection before the Chinese or Asian epileptics take carbamazepine and phenytoin sodium.

The invention relates to a primer pair and kit for detecting VKORC1 (vitamin K epoxide reductase complex subunit 1) genotyping by pyrosequencing, belonging to the technical field of in vitro nucleic acid detection. The primer pair comprises VKORC1 G1639A and VKORC1 C1173T forward amplification primers, VKORC1 G1639A and VKORC1 C1173T reverse amplification primers and VKORC1 G1639A and VKORC1 C1173T sequencing primers, wherein 5' terminals of the VKORC1 G1639A forward amplification primer and the VKORC1 C1173T reverse amplification primer are respectively subjected to biotin labelling. The kit comprises the amplification primers, a PCR (polymerase chain reaction) liquid 1, a PCR liquid 2, the sequencing primers, uracil DNA (deoxyribonucleic acid)glycosylase and Taq polymerase. The kit provided by the invention has the advantages of accurate detection results, high specificity, short detection period, simplicity in operation, capability of effectively meeting the requirements of clinical examination, capability of monitoring the reaction process in real time, short reaction time, sequencing of PCR products on a pyrosequencing instrument after the PCR products are simply treated, high throughput sample detection and higher sensitivity than gold standard methods, namely capillary electrophoresis sequencing methods.

The invention provides a nonenzymatic biosensor for detecting activity of uracil-DNA glycosylase. The nonenzymatic biosensor comprises a UDG template strand 1, a silver nanocluster DNA template strand2, and a DNA strand 3 rich in cytosine. A fluorescence method is used for detecting the activity of the uracil-DNA glycosylase, the stimulation wave lenagth of fluoroscopic examination is 570nm, thelength of transmitted waves is 630nm, and the detection wave band is 590-720nm. The biosensor provided by the invention is good in specificity, high in sensitivity, mild in reaction condition and highin reaction speed. Because the reaction process is free from addition of external enzymes, and through silver nanocluster fluoroscopic examination, the biosensor is simple to operate, low in reactionrequirements, short in detection cycle, stable in properties, and good in repeatability, and is suitable for detection of UDG in the field of medical health.

The invention provides a sequencing primer pair for qualitatively detecting human BRAF V600E gene mutation and a kit thereof, belonging to the field of detection for in-vitro nucleic acid. The kit comprises a uracil DNA (deoxyribonucleic acid) glycosylase, a Taq polymerase, PCR (polymerase chain reaction) solution, a PCR amplification primer, a pyrosequencing primer and a positive reference substance. The kit provided by the invention is high in sensitivity, good in specificity, capable of sequencing a PCR product on a pyrosequencer after performing a simple treatment, simple and convenient in operation, short in reaction time, higher in sensitivity compared with golden standard-capillary electrophoresis sequencing, and more suitable for mutation analysis.

Described are methods of detecting modified nucleotide bases in a DNA sample using specific DNA glycosylases to excise target modified bases. DNA molecules are then labeled using a DNA polymerase lacking 3′→5′ exo-nuclease activity and strand displacement activity. The methods can be used to detect epigenetic changes and DNA damage. Provided are methods for diagnosing a disease or condition, determining risk of a disease or condition, identifying appropriate treatment, monitoring effectiveness of treatment, and monitoring side effects of treatment in subjects based on detection of modified bases. Also provided are methods for determining environmental exposure, or an environmental exposure time, of a biological sample containing DNA. Also provided are kits, systems, and devices for performing the described methods.

The invention belongs to the technical field of biological detection and molecular biology, and particularly relates to a DTNS-mediated method for detecting the activity of 8-OGDNA glycosylase. Under the action of 8-OG DNA glycosylase, the structure of the DTNS is changed from an open state to a closed state, so that the distance between a Cy3 donor and a Cy5 receptor is close, and efficient FRET is initiated. By utilizing the method, the activity of extracellular 8-OG DNA glycosylase can be sensitively and selectively detected, and false positive signals generated by nuclease degradation can be prevented based on an FRET signal output mode, and accurate imaging in living cells is facilitated.

The invention belongs to the technical field of molecular detection, and particularly relates to a fluorescent chemical sensor for simultaneously detecting multiple DNA glycosylase as well as a detection method and application of the fluorescent chemical sensor. The fluorescent chemical sensor comprises a double-stranded DNA substrate, two or more than two DNA glycosylase recognition sequences and a reporter probe, wherein the double-stranded DNA substrate comprises a recognition sequence for recognizing one or more DNA glycosylase; the 5' tail ends of the two chains of the double-chain DNA substrate are respectively modified with fluorescence quenching groups; the reporter probe is modified by a fluorophore; the base sequence connected with the fluorescence quenching group is hybridized with the reporter probe to form a base pair. According to the invention, the signal amplification technology based on the fluorescence chemical sensor can realize ultra-sensitive detection of various DNA glycosylase.

Provided herein are methods and compositions for inducing chemical senescence in neurons and methods of using chemically induced senescent neurons for modeling neurodegenerative disease and drug discovery. The methods include contacting human neurons with a culture medium comprising an inhibitor of DNA glycosylase 1, an autophagy inhibitor, and an HIV protease inhibitor to obtain an in vitro population of senescent neurons within about 4 days. When the neurons are obtained from a patient having a neurodegenerative disease, chemically induced senescent neurons obtained by these methods recapitulate cellular and subcellular phenotypes observed in individuals with the neurodegenerative disease.

The invention relates to a primer pair and a reagent kit for detecting hepatitis B canceration susceptibility gene polymorphism, and belongs to the technical field of in vitro nucleic acid detection.The primer pair comprises a forward amplification primer, a reverse amplification primer and a sequencing primer, wherein the 5'-end of the reverse amplification primer is labeled with biotin. The kitcomprises a PCR (Polymerase Chain Reaction) solution containing a forward amplification primer, reverse amplification primer, a sequencing primer, uracil DNA (Deoxyribonucleic Acid) glycosylase, Taqpolymerase and a positive control product. The kit provided by the invention has the advantages of accurate detection result, high specificity, short detection period, and simple operation, and the clinical inspection requirements can be effectively met; in addition, the kit also has the advantages of real-time monitoring of reaction progress, and short reaction time, the PCR product can be used for sequencing on a pyrosequencing instrument after being simply treated, the sensitivity is higher than that of a gold standard method, i.e., a capillary electrophoresis sequencing method, and the kitis more applicable to mutation analysis.

The invention belongs to the technical field of biology, and particularly relates to an 8-oxoguanine DNA glycosylase determination method based on a background signal inhibition probe, a kit and an application thereof. According to the present invention, the selectivity of Lambda exonuclease on the substrate structure and the recognition cutting effect of 8-oxo guanine DNA glycosylase on 8-oxo guanine are utilized, and the inhibition effect of the background signal inhibition probe on the lambda exo side activity is combined so as to construct the new 8-oxo guanine DNA glycosylase fluorescence determination method. The method specifically comprises the following steps: mixing a lambda exo buffer solution, a reporter probe sequence containing a fluorophore and a quenching group, and a background signal inhibition probe sequence containing 8-oxo guanine; heating the mixed solution to 85 DEG C, then gradually cooling to 37 DEG C, and adding a sample to be detected; incubating at 37 DEG C; and after incubation is finished, adding lambda exo, carrying out enzyme digestion reaction, and then carrying out fluorescence detection.