52 results about "Methyl cytosine" patented technology

The present invention provides for novel methods for regulating and detecting the cytosine methylation status of DNA. The invention is based upon identification of a novel and surprising catalytic activity for the family of TET proteins, namely TET1, TET2, TET3, and CXXC4. The novel activity is related to the enzymes being capable of converting the cytosine nucleotide 5-methylcytosine into 5-hydroxymethylcytosine by hydroxylation.

The invention belongs to the technical field of biological detection, and particularly relates to a bisulfite-free detection kit for specific site 5-methylcytosine and a detection method thereof. According to the method, a TAPbeta treatment method is adopted, 5hmC is sealed through beta-GT, 5mC oxidation is carried out through TET, and 5mC is reduced into dihydrouracil (DHU) through pyridine borane, so that the apparent difference between 5mC and C and 5hmC is converted into single nucleotide polymorphism difference. In addition, a flow microsphere technology based on PNA-assisted click chemical connection starting is established to detect DNA methylation specific sites. A PNA clip is innovatively introduced into click chemical connection, and non-specific connection is inhibited. Target sequence enrichment, signal amplification and signal acquisition are carried out through surface-functionalized magnetic nanoparticles, and the detection sensitivity is improved to fM. The invention provides a new tool for high-specificity and high-sensitivity epigenetic marker detection.

The invention provides a method for the analysis of a biological sample to determine multiple types of information therefrom in a streamlined, combined workflow, where all information is obtained in a sequencing-based analysis. The information includes the presence and concentration of specific plasma proteins in a blood sample: the number, location, and types of histone modifications associated with cell-free DNA obtained from the same sample: the sequence of cfRNA and cfDNA in the cell-free DNA sample; and epigenetic information pertaining to the cell-free DNA, such as hydroxy methylation and methylation profiles, i.e., the distribution of 5-hydroxymethylcytosine (5hmC) and 5-methylcy tosine (5mC) residues, respectively. The invention additionally pertains to a classical sequencing-based method for analyzing a biological sample to determine one or more non-classical sequence features of the sample. Compositions, kits, and related methods are also provided, including an embodiment in which truncated sequencing adapters are used in conjunction with barcoded PCR primers.

To provide an antisense oligonucleotide for preventing, delaying, and / or treating a human cis-element repeat instability-associated genetic disorder.SOLUTION: A medicament comprises, as an active ingredient, an oligonucleotide, wherein the oligonucleotide comprises 2'-O-methyl RNA nucleotide residues, has a backbone with at least one phosphate moiety being replaced by a phosphorothioate moiety, and comprises or consists of a repetitive nucleotide unit (XYG)m, wherein m is an integer from 4 to 12, each X is C or 5-methylcytosine, each Y is U or 5-methyluracil, and at least one X is 5-methylcytosine.SELECTED DRAWING: None

This disclosure provides methods for cost-effective bisulfite-free identification in DNA, including whole genomic DNA, of the locations of one or more of 5-methylcytosine, 5-hydroxymethylcytosine, 5-carboxylcytosine and 5-formylcytosine. The methods described herein are based on the conversion of modified cytosine (5mC, 5hmC, 5fC, 5caC) to dihydrouracil (DHU), for example by TET-assisted pyridine borane treatment, followed by endonuclease cleavage of the DHU, and identification of the cleavage site, which corresponds to the location of the modified cytosine.

A method for detecting a methylated genomic locus is provided. In certain embodiments, the method comprises: a) treating a nucleic acid sample that contains both unmethylated and methylated copies of a genomic locus with an agent that modifies cytosine to uracil to produce a treated nucleic acid; b) amplifying a product from the treated nucleic acid using a first primer and a second primer, wherein the first primer hybridizes to a site in the locus that contain methylcytosines and the amplifying preferentially amplifies the methylated copies of the genomic locus, to produce an amplified sample; and c) detecting the presence of amplified methylated copies of the genomic locus in the amplified sample using a flap assay that employs an invasive oligonucleotide having a 3′ terminal G or C nucleotide that corresponds to a site of methylation in the genomic locus.

The invention relates to the technical field of biology, in particular to 5-hydroxymethylcytosine and application of a prognosis model of 5-hydroxymethylcytosine in nasopharynx cancer prognosis evaluation. According to the method, the prognosis scoring model capable of layering the outcome of the patient is constructed, and the model shows strong distinguishing performance in a training set and a verification set. When it is integrated into a prognostic model together with tumor staging and EBV status, the 5hmC score results in higher calibration accuracy and net clinical benefit in decision curve analysis. According to the findings, a 5hmC prognosis model is determined as a noninvasive marker rich in biological information, can be used for prognosis risk layering of NPC patients, and has potential significance for individualized management and improvement of outcome prediction.

The present invention discloses a method for detecting 5-hydroxymethylcytosine in the whole genome and its application, which relates to the technical field of biochemical detection. The method comprises the following steps: 1: Blocking the 3'-OH end of the DNA to be detected; 2: Preparation of glycosylated 5hmC DNA; 3: TdT-mediated DNA amplification; 4: Fluorescence measurement. The present invention constructs a homogeneous, template-independent signal amplification method for detecting 5-hydroxymethylcytosine in the whole genome. This method is based on specific enzyme-catalyzed reactions and multiple signal amplification strategies, and has the characteristics of high sensitivity, good specificity, no need for separation steps, sequence-independent amplification, and the ability to detect 5hmC at any position in the whole genome. Moreover, the present invention can achieve accurate and sensitive detection of actual clinical samples.

Methods, compositions, and systems are provided for detecting 5-methylcytosine modifications in nucleic acids, particularly DNA samples. One or more templates are sequenced using single molecule real-time sequencing. A feature vector is generated using the particular feature. In some aspects, feature vectors with reduced feature sets are provided, and these feature vectors are input into a deep learning model.

A method of determining percentages of twelve blood cell subtypes in a blood specimen, includes steps of: (a) obtaining genomic DNA from the blood specimen of a human subject, (b) observing unmethylated CG loci, in the genomic DNA of the human subject, wherein said observing includes performing a bisulfate conversion process on the genomic DNA of the human subject so that cytosine residues in the genomic DNA of the human subject are transformed to uracil, while 5-methylcytosine residues in the genomic DNA of the human subject are not transformed to uracil, (c) comparing the unmethylated CG loci observed in (b) to unmethylated CG loci observed in genomic DNA collected from a reference group of human individuals and (d) correlating the unmethylated CG loci observed in (b) with the unmethylated CG loci observed in the reference group of human individuals to determine the percentage of the twelve blood cell subtypes in the blood specimen.

Disclosed herein are aqueous compositions that include a pyridine complex and a polynucleotide. The compositions can be used to detect methylcytosine and / or hydroxymethylcytosine in the polynucleotide. In some examples, the compositions may be used as part of a TET-assisted borane sequencing workflow.

Protein complexes including a cytidine deaminase and a helicase. In some embodiments, the cytidine deaminase is an altered cytidine deaminase. In some embodiments, the protein complex converts 5 methylcytosine to thymine. Kits, compositions, and methods of use for the protein complexes including a cytidine deaminase and a helicase are also described.

The invention includes improved methods and compositions for detecting methylation in nucleic acids. In particular, the disclosure is directed to methods of converting 5-hydroxymethylcytosine (5hmC) and / or 5-methylcytosine (5mC) to Thymine (T). In addition, the disclosure is also directed to methods of detecting 5hmC and / or 5mC in a sample.

The present invention relates to methods and kits for the detection of 5-hydroxymethylcytosine (5hmC) and / or 5-methylcytosine (5meC). In some embodiments, the present invention relates to methods and kits for detection of 5hmC and / or 5meC in nucleic acid (e.g., DNA, RNA). In some embodiments, the present invention relates to detection of 5hmC in genomic DNA, e.g., mammalian genomic DNA

The disclosed technology relates to a method of analyzing DNA from a biological sample. In one aspect, the method comprises obtaining a sample a sample from a subject, the sample comprising DNA that comprises double-stranded DNA (dsDNA) suspected of having one or more thymine lesions formed by deamination of 5-methylcytosines, wherein the thymine lesions form base pair mismatches in the dsDNA, contacting the sample with a nuclease that digests dsDNA at base pair mismatches; harvesting DNA from the nuclease-treated sample; and performing an analysis on undigested DNA harvested from the nuclease-treated sample.

The invention discloses a device for screening patients with minimal lesion type nephropathy and application of the device in screening the patients with minimal lesion type nephropathy. The device for screening the patients with minimal lesion type nephropathy comprises a substance for detecting 5-methylcytosine modification of miR-511-5p, or comprises a substance for detecting the expression quantity of an ENPP4 gene, a substance for detecting the expression quantity of an ENTPD1 gene, a substance for detecting the expression quantity of a DNMT1 gene and a substance for detecting the expression quantity of an RND3 gene. The invention finds that m5C modification of miR-511-5p in an MCD patient is down-regulated, key genes ENPP4, ENTPD1 and DMNT1 in peripheral blood of the MCD patient are further screened, expression of the key genes is down-regulated, and miR-511-5p affects lipid metabolism of renal tissues by down-regulating RND3. The device for screening the patients with the minimal lesion type nephropathy can be used for screening the patients with the minimal lesion type nephropathy.

The present disclosure is concerned with modified proteins, methods, compositions, and kits for mapping of methylation status of nucleic acids, including 5-methylcytosine. In some embodiments, the proteins are altered cytidine deaminases, such as altered APOBEC3A proteins. In some embodiments, the proteins selectively act on cytosines and have significantly reduced activity on modified cytosines, such as 5-methylcytosine.

The invention provides a targeted ALKBH2 small-molecule inhibitor and application thereof, and belongs to the field of medicinal chemistry. The structure of the small-molecule inhibitor is shown as a formula I. The compound provided by the invention has good affinity activity on ALKBH2, can competitively inhibit recognition of the ALKBH2 on a substrate in a targeted manner, and specifically targets ALKBH2 instead of other AlkB family proteins. The compound provided by the invention can be used for effectively targeting ALKBH2 in cells and increasing the modification abundance of N3-methylcytosine (m3C) in the cells. The compound provided by the invention not only can effectively inhibit proliferation of brain glioma cells, but also can effectively inhibit migration of the brain glioma cells. The compound provided by the invention can be used for preparing ALKBH2 inhibitors and medicines for preventing or treating ALKBH2 expression abnormity related diseases (such as brain tumor, non-small cell lung cancer, rectal cancer, bladder cancer and gastric cancer), and has a wide application prospect.

The present invention provides oligonucleotides useful for the treatment of Duchenne muscular dystrophy or Becker muscular dystrophy, compositions containing oligonucleotides, and methods for prevention, treatment, and / or delay. [Solution] An oligonucleotide is provided that comprises a 2'-O-methylRNA monomer and a phosphorothioate skeleton, and also comprises 5-methyluracil and / or 5-methylcytosine and / or a 2,6-diaminopurine base.

The application discloses a data analysis method in the field of epigenetics: a quantitative analysis method of RNA 5-methylcytosine modification on splicing isomers. The method can solve the problem of how to quantitatively analyze RNA 5-methylcytosine modification on the level of gene splicing isomers, and the technical points include: converting the data set generated by the RNA-BSseq sequencing technology into a virtual RNA-Seq data set and marking the data in which RNA 5-methylcytosine modification occurs; estimating the expression amount of the splicing isomer in the virtual data set, and calculating the mean and variance thereof by using the bootstrap technology; estimating the RNA 5-methylcytosine modification level of the splicing isomer, and calculating the mean and variance of the RNA 5-methylcytosine modification level of the splicing isomer by using the Delta technology; and based on the normal distribution, testing whether there is a significant difference in the estimated RNA 5-methylcytosine modification level of the splicing isomer under different conditions. The application helps to reveal the regulation mechanism of RNA 5-methylcytosine modification in physiological processes and pathological changes.

The present disclosure relates generally to the enzymatic conversion of methylated nucleic acids in order to differentiate methylated cytosine from unmethylated cytosine in DNA, and more particularly to improved methods and compositions for enzymatic methylation sequencing. In one aspect, various compositions and methods for improving the recovery of methylation signals are provided. The methods include one or more of a nick repair step, methylation signal restoration, use of a modified methylcytosine nucleic acid adapter, and use of a helicase, an ssDNA binding protein, an engineered DNA ligase, or a combination thereof.

The present disclosure is concerned with modified proteins, methods, compositions, and kits for mapping of methylation status of nucleic acids, including 5-methylcytosine and 5-hydroxymethyl cytosine (5hmC). The proteins selectively act on certain modified cytosines of target nucleic acids and include one or more substitution mutations that enhance the selectivity of the proteins for certain modified cytosines, enhance the stability of the proteins, or enhance both selectivity and stability. Also provided are compositions and kits that include one or more of the proteins and methods for using one or more of the proteins.

The invention relates to the technical field of biochemical detection, in particular to a sample pretreatment method, a method for detecting the ratio of 5-methylcytosine to cytosine and a kit. The sample pretreatment method comprises the following steps: providing a sample containing substances to be detected, wherein the substances to be detected comprise 5-methylcytosine and cytosine; the method comprises the following steps: mixing magnetic beads containing sulfonic acid groups with a sample, carrying out adsorption treatment, and then carrying out leaching treatment by using an alcoholic solution to obtain a to-be-detected substance-magnetic bead compound; eluting the to-be-detected substance-magnetic bead compound by using the eluent to obtain a to-be-detected sample; the eluent comprises at least one of a formic acid-alcohol solution and an ammonia water-alcohol solution. The sample pretreatment method is simple in process and low in cost, the to-be-detected sample containing the 5-methylcytosine and the cytosine can be efficiently obtained, the ratio of the 5-methylcytosine to the cytosine can be accurately quantified in a high-sensitivity mode on the basis of the electrospray high-efficiency ion mobility spectrometry technology subsequently, and the method has good application prospects.

To provide compositions and methods for detecting gastrointestinal pathogens.SOLUTION: Compositions, methods, kits, and uses are provided for detecting the presence of Salmonella, Shigella, Campylobacter, and Shigatoxigenic Eschelichia coli (STEC) in samples. In some embodiments, the compositions, methods, kits, and uses comprise one or more oligonucleotides or the use of one or more oligonucleotides. In some embodiments, the method is carried out as a multiplex assay with separate primers / probes for C.jejuni and C.coli. Some of the oligonucleotides contain inosine and / or contain 5-methylcytosine substituted nucleotides. Described is the use of cyclodextrin and / or Polysorbate 20 in the amplification reactions.SELECTED DRAWING: None

The invention discloses a stomach cancer early screening ELISA kit and an early screening detection method thereof, and relates to the technical field of in vitro diagnosis. The kit comprises the following components: a pre-coated 96-well plate which is fixedly provided with methylated DNA fragments of an anti-5-methylcytosine monoclonal antibody (with the clone number of 33D3), cg07160992, cg13516541, cg02956093 and cg21496913; the detection assembly comprises a biotinylation detection probe and a streptavidin-HRP (horse radish peroxidase) conjugate; the pretreatment assembly comprises a hydrosulfite conversion reagent and MBD2 protein modified magnetic beads. According to the kit, through a double-index combined interpretation model, the sensitivity to early gastric cancer reaches 86%, the blank of non-invasive detection of stage I gastric cancer is filled, the total sensitivity is 90.5%, the international top-level cancer early screening product standard is met, operation is standardized, special instruments are not needed, and the kit has high clinical application value.

The application provides a nucleic acid aptamer-drug conjugate, a preparation method thereof and application thereof in preparing an anti-tumor drug, and belongs to the technical field of biological medicines. The nucleic acid aptamer-drug conjugate comprises a nucleotide sequence, an HG9 nucleic acid aptamer as shown in SEQ ID NO:1 and an extended chain at the end of the HG9 nucleic acid aptamer; the extended chain is loaded with 5-hydroxymethylcytosine and / or 5-formylcytosine. The application further provides a preparation method of the nucleic acid aptamer-drug conjugate. By means of PCR amplification technology, an epigenetic modification nucleoside with cytotoxicity is introduced into a nucleic acid aptamer with specific targeting capability, the nucleic acid aptamer-drug conjugate constructed has the binding capability to target cells, and can release the modified nucleoside in the cells, cause DNA damage and significantly inhibit the activity of the target cells. The method is simple in operation and high in efficiency, and the obtained conjugate has potential value in drug development.

Disclosed herein are compositions for preparing a target nucleic acid molecule comprising one or more 5-formylcytosine bases or adducts of 5-formylcytosine. Also disclosed herein are methods of efficiently synthesizing a nucleic acid molecule comprising one or more 5-formylcytosine bases from a target nucleic acid molecule comprising one or more 5-hydroxymethylcytosine bases. The disclosure also provides methods of detecting epigenetic modifications in a target nucleic acid molecule, such as those characterized by the methylation of cytosine at position 5 (e.g., 5-methylcytosine, 5-hydroxymethylcytosine).

The invention provides a nucleic acid aptamer-drug conjugate as well as a preparation method and application thereof in preparation of anti-tumor drugs, and belongs to the technical field of biological medicines. The nucleic acid aptamer-drug conjugate comprises an HG9 nucleic acid aptamer with a nucleotide sequence as shown in SEQ ID NO: 1 and an extension chain at the end part of the HG9 nucleic acid aptamer; 5-hydroxymethylcytosine and / or 5-formyl cytosine are / is loaded in the extension chain. The invention further provides a preparation method of the nucleic acid aptamer-drug conjugate, apparent modified nucleoside with cytotoxicity is introduced into the nucleic acid aptamer with specific targeting capacity through a PCR amplification technology, and the constructed nucleic acid aptamer-drug conjugate has the binding capacity to target cells, can release the modified nucleoside in the cells, and can be used for preparing the nucleic acid aptamer-drug conjugate. DNA damage is triggered; and the activity of target cells is remarkably inhibited. The method is simple and convenient to operate and high in efficiency, and the obtained conjugate has potential drug development value.

Methods for inhibiting respiratory viruses (i.e., viruses associated with respiratory conditions, e.g., influenza A, influenza B, RSV, and the like) are provided. Aspects of the methods include contacting a sample comprising viral RNA (vRNA) having a target motif with an effective amount of an agent that specifically binds to the target motif to inhibit the respiratory virus. Methods for treating or preventing respiratory virus infection in a subject are also provided. Pharmaceutical compounds and compositions comprising an oligonucleotide sequence complementary to a target region of the vRNA used in the methods are also provided. The oligonucleotides provided may include a 5-methylcytosine.

The present invention discloses a use of 5-hydroxymethylcytosine, and provides an application of 5-hydroxymethylcytosine in the diagnosis of systemic lupus erythematosus or in the preparation of a product for the diagnosis of systemic lupus erythematosus; the hydroxymethylation biomarker is one or more loci of ETS1. The present invention has the characteristics of strong sensitivity and high specificity, and can be applied to the auxiliary diagnosis of clinical systemic lupus erythematosus patients. By combining other clinical symptoms, signs and laboratory indicators, the diagnosis rate of early systemic lupus erythematosus can be improved, and clinicians can be assisted in formulating treatment plans in a timely manner, improving the diagnosis and treatment level of SLE and improving patient prognosis. In addition, the present invention also discloses a kit for diagnosing the above-mentioned biomarkers of SLE patients, which is intended to comprehensively improve the ability of clinicians to perform early diagnosis and preemptive intervention treatment on SLE patients.