56 results about "Methylation analysis" patented technology

Methods, compositions and kits are provided to amplify an amount of genomic methylated DNA that can be subsequently analyzed and / or sequenced. It has particular use with small amounts of DNA, including, but not limited to cell free DNA samples. In some embodiments, the ratio of polymerase and methyltransferase is controlled in order to provide maximum yields. In some embodiments, a dual primase / polymerase is used.

The present invention discloses an in vitro method for obtaining DNA methylation biomarkers as exquisite DNA methylation positions in the human genome (i.e., CGIDs) that predict cervical cancer especially at as yet inaccessible early stages by examining progression of “categorical” DNA methylation alterations in three stages of premalignant lesions (cervical intraepithelial neoplasia (CIN)), progressing from CIN1 to CIN3. The present invention discloses combinations of CGIDs for detecting with high specificity and sensitivity cervical cancer by measuring their DNA methylation status and deriving a “methylation score”, which is useful as a biomarker for cervical cancer. Also disclosed are kits for predicting cervical cancer using such CGIDs using multiplexed next generation sequencing methylation assays, pyrosequencing assays and methylation specific PCR. The DNA methylation markers (CGIDs) described in the present invention are useful for cervical screening and early detection of cervical cancer by any person skilled in the art to detect cervical cancer.

Described herein are methods of preparing dual-indexed nucleic acid libraries for methylation profiling using bisulfite conversion sequencing. In various embodiments, the methods use a two-step indexing process to tag bisulfite-treated DNA with unique molecular identifiers (UMIs).

Described herein are dual-indexed nucleic acid libraries for methylation profiling using bisulfite conversion sequencing and methods of preparing same..

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.

Polysaccharide is a main bioactive component in rhizoma polygonati. According to the research, the rhizoma polygonati polysaccharide PSPF is obtained by adopting DEAE-sepharose FF and Sephadex LH-20 column chromatography, and the rhizoma polygonati polysaccharide PSPF is characterized through gel permeation chromatography, monosaccharide analysis, FT-IR (Fourier transform infrared spectroscopy), NMR (nuclear magnetic resonance) and GC-MS (gas chromatography-mass spectrometry) methylation analysis. And the weight-average molecular weight of the PSPF is 3592 Da. The result shows that the PSPF is mainly composed of fructose, and the fructose has a main chain of-> 1)-beta-D-fructofuranose-(2) and a side chain of-> 6)-beta-D-fructofuranose-(2). The PSPF can reduce the blood fat of an acute hyperlipidemia model, the serum HDL-C is obviously increased, and the oxidation and decomposition of fatty acid can be promoted, which is related to the up-regulation of AMPK and CPT-1 and the low expression of SREBP-1c, ACC and FAS; therefore, the PSPF is an apparent food polysaccharide for controlling lipid.

The present disclosure provides methods for generating mbias corrected sequence fragment data comprising correcting one or more strand displacement mbias instances in one or more nucleic acid fragments from a sample obtained from a subject, performing a methylation analysis, and correcting one or more end-repair bias instances in sequence fragment data from the methylation analysis.

A method is provided for probabilistically assigning a tissue of origin to a nucleic acid in a sample, e.g., DNA in a cell-free fluid sample obtained from a human subject. A hydroxymethylation profile is generated for the sample DNA and then compared across a reference data set of hydroxymethylation profile vectors, where each hydroxymethylation profile vector identifies the hydroxymethylation profile at a specific reference locus, the tissue-specific gene associated with the reference locus, and the tissue with which the gene and reference locus are associated. A tissue of origin can be probabilistically assigned to the sample nucleic acid using the results of the comparison. Other methods of use are also provided.

This invention discloses a method for constructing a gene methylation diagnostic model for differentiated thyroid cancer (PTC). The method includes the following steps: S1, obtaining a test sample; S2, extracting and storing DNA from the test sample; S3, performing methylation analysis; S4, establishing a diagnostic model based on DNA methylation, where the model shows that patients with cg03596178, cg06033721, cg06688989, cg07209244, cg07485775, cg14484681, cg19979108, and cg20943461 are more likely to experience disease progression and are considered risk factors; S5, cross-validating and evaluating the performance to obtain the gene methylation diagnostic model. This model, by detecting and statistically analyzing specific methylation sites, can differentiate patients with recurrent or metastatic characteristics in differentiated thyroid cancer. The relevant model can assist in the clinical diagnosis and follow-up of thyroid patients.

The invention discloses a gene screening method influencing drought adaptability of Hu sheep, and relates to the technical field of molecular breeding. The method comprises the following steps: step 1, selection and treatment of experimental sheep; step 2, transcriptome analysis; step 3, carrying out methylation analysis on the whole genome; and step 4, carrying out transcriptome and whole genome methylation conjoint analysis. The LOC121817420 gene is mined through a transcriptome and whole genome methylation method, the expression quantity of the gene in Hu sheep whole blood in two regions is detected through a real-time fluorescence quantification method, the expression quantity of the gene in Hu sheep in the Hetian region of Xinjiang is high, and it is proved that the gene is one of main candidate genes, different from Hu sheep in the Taihu Lake basin, of the Hu sheep in Xinjiang.

The invention discloses a gene level DNA methylation, transcriptome and proteome conjoint analysis method based on a snkemake framework, which comprises the following steps: preprocessing original data of DNA methylation and / or transcriptome and / or proteome, analyzing gene methylation, analyzing transcriptome and / or proteome, and analyzing the transcriptome and proteome. Based on gene methylation analysis data, the preprocessed transcriptome data and the preprocessed proteome data, correlation analysis and / or difference intersection analysis and / or enrichment pathway conjoint analysis are / is carried out, finally analysis results are sorted, and a visual report is generated. The method has comprehensive results, and relates to quantification, difference analysis and pathway enrichment analysis of gene methylation, and correlation analysis, difference intersection analysis and enrichment pathway joint analysis of integrated DNA methylation and / or transcriptome and / or proteome data; automatic arrangement, verification, visualization and report generation of analysis results are realized; all operation steps can be traced, and corresponding analysis log records are generated. The invention further discloses a related system and application.

The invention relates to the technical field of biological pharmacy, in particular to the field of IPC C08B37, and more particularly relates to a structure identification method of bletilla striata neutral polysaccharide. Comprising the following steps: step 1, sequentially extracting, separating and purifying rhizoma bletillae crude polysaccharide to obtain rhizoma bletillae polysaccharide; step 2, pretreating the rhizoma bletillae polysaccharide, and performing homogeneity and molecular weight determination; 3, determining the content of total sugar in the rhizoma bletillae polysaccharide by adopting a phenol-sulfuric acid method, determining the chemical composition in the rhizoma bletillae polysaccharide by adopting a trifluoroacetic acid method in combination with gas chromatography-mass spectrometry, determining the content of uronic acid in the rhizoma bletillae polysaccharide component by adopting an m-hydroxybiphenyl method, and determining the content of protein in the rhizoma bletillae polysaccharide by adopting a Bio-rad protein quantitative method; step 4, carrying out methylation analysis on the bletilla striata polysaccharide; and step 5, analyzing monosaccharide components and bonding modes of the bletilla striata polysaccharide by using a nuclear magnetic resonance technology.

The present disclosure provides a method for methylation analysis of genomic fragments.

In an aspect, the present disclosure provides a method comprising (a) providing a cell-free deoxyribonucleic acid (cfDNA) sample derived from a subject; and (b) sequencing the cfDNA sample or a derivative thereof to determine a methylation pattern or a methylation level of DNA molecules of the cfDNA sample.

The present invention relates to a real-time polymerase chain reaction method for determining a degree and pattern of DNA methylation at a specific DNA sequence site and to a use thereof. More specifically, the present invention relates to a method for determining a degree and pattern of methylation by determining respective Ct values obtained from performing qPCR in two buffer systems (buffer 1 and buffer 2), in which differences occur depending on the presence or absence of methylation, using DNA obtained from a biological sample, calculating a difference between the Ct values of the two buffer systems, and more specifically, calculating a relative ratio of copy number calculation values obtained from tests in the two buffer systems, and to a use thereof.

Methods for mitigation of methylation bias during preparation of sequencing libraries are described. The methods may comprise, e.g., extracting DNA fragments from a sample; performing at least one of: (i) a first end repair reaction, where the first end repair reaction comprises the use of a non-strand-displacing DNA polymerase; or (ii) a second end repair reaction, where the second end repair reaction comprises the use of a chain termination mechanism; performing at least one of: (iii) a tailing reaction to add an overhanging polynucleotide strand to the end-repaired DNA fragments, where the tailing reaction comprises the use of a single deoxynucleotide triphosphate (dNTP); or (iv) a nick / gap repair reaction using a DNA ligase to fill in single- stranded nicks or gaps in the end-repaired DNA fragments, to generate a plurality of modified DNA fragments; and performing methylation analysis of nucleic acid molecules derived from the plurality of modified DNA fragments.

The application discloses a nucleic acid sequence construction method for cfDNA multiplex gene methylation detection and application thereof. The construction method comprises the following steps: using a methylation-sensitive restriction endonuclease to perform enzyme cutting on to-be-detected cfDNA to obtain enzyme cutting fragments, using a ligase to connect an adapter to a nick end of the enzyme cutting fragments to form an adapter product; and using the adapter product as a template to perform PCR amplification to construct a recombined nucleic acid sequence for target gene methylation site detection. The construction method can be used for cfDNA level methylation qPCR analysis and low-abundance sample quantitative analysis, has small sample demand, has a wide methylation analysis coverage, and has high methylation enrichment efficiency.

The invention discloses ulva pertusa polysaccharide UPP and ulva pertusa homogeneous polysaccharide UPP-5 as well as a preparation method and application thereof. The single-component polysaccharide UPP-5 is obtained by adopting DEAE-52 anion exchange chromatography in combination with ultrafiltration purification. The structure of the UPP-5 is systematically characterized by high performance gel permeation chromatography, infrared spectroscopy, methylation analysis, nuclear magnetic resonance and other methods, and the result shows that the UPP-5 is acidic heteropolysaccharide with a novel structure. Based on an AOM / DSS induced colorectal cancer mouse model, the effect of the mouse model is evaluated from the aspects of disease activity index, colon length, tumor number, histological observation, gene and protein expression and the like. Results show that the UPP can significantly improve inflammation and pathological states and reduce the incidence of tumors, and has the potential of preventing and treating colorectal cancer. The invention provides a new thought and theoretical basis for structural research and medical function development of algal polysaccharides.

The invention discloses a DNA and RNA methylation analysis method for a single cell or a small amount of cells. The method comprises the following steps: A, directly using a single cell or a small amount of mammalian cells; b, the purpose of breaking cells is achieved by directly using high temperature or protease treatment; c, treating the non-methylated DNA by using a methylation-sensitive incision enzyme; e, directly treating the sample by using hydrosulfite to convert unmethylated cytosine (C) in the DNA and RNA of the sample into uracil (U); f, simultaneously carrying out methylation analysis on DNA and RNA from the same cell source; g, DNA and RNA methylation results of a single cell source can be obtained at the same time. According to the method, DNA and RNA in a single cell are effectively utilized, the method is easy to implement, stable in result, high in detection sensitivity, more sufficient in information and simple and convenient to operate, and the limitation of the DNA amount of the single cell in DNA methylation analysis is greatly improved.

Systems and methods of the disclosure may include a computer-implemented method, the computer-implemented method including: receiving, at a computer system, nucleic acid sequencing data derived from a methylation assay performed on a biological sample associated with at least one subject; computing, using a processor associated with the computer system, a beta value matrix based on the nucleic acid sequencing data, wherein the beta value matrix comprises one or more missing beta values; addressing, using the processor, the one or more missing beta values in the beta value matrix using a missing beta value completion approach; identifying, using the processor, one or more principal components in the completed beta value matrix; and training, using the one or more principal components in combination with a predetermined set of clinical variables, a classifier to predict a survival outcome for a target subject associated with a disease type.

This invention discloses a method for constructing a gene methylation prognostic assessment model for differentiated thyroid cancer, characterized by the following steps: S1, obtaining a test sample; S2, extracting and storing DNA from the test sample; S3, performing methylation analysis; S4, constructing a prognostic classification model based on DNA methylation, calculating the risk value of the prognostic model as Risk Score = 0.15411928*cg03190661 - 0.10405129*cg15676916 + 0.06108015; S5, cross-validating to evaluate performance and obtain the gene methylation prognostic assessment model. This model uses the Risk Score to assess the prognosis of differentiated thyroid cancer: low-risk group, normal follow-up is recommended; medium-risk group, follow-up time can be reduced to half; high-risk group, close follow-up is required.

The present disclosure provides methods for detecting a presence or an absence of a liver disease in a subject. An exemplary method includes assaying a bodily sample obtained or derived from the subject to determine quantitative measures of a set of liver-specific methylated biomarkers in a nucleic acid from the bodily sample, computer processing the set of liver-specific methylated biomarkers using a trained machine learning algorithm or compared against a reference, and detecting the presence or the absence of the liver disease in the subject, based at least in part on the computer processing.

The invention discloses a method for identifying and analyzing m6A methylation modification sites of dengue viruses based on MeRIP-seq, which comprises the following steps of: sequencing by combining a specific m6A antibody with m6A modified RNA (Ribonucleic Acid), and mapping reads onto a reference genome by using HISAT2 and other software packages to determine 6 m6A peaks of DENV-I type and 5 m6A peaks of DENV-II type. The method can identify and locate m6A modification, can be used for m6A methylation analysis of a whole transcriptome level, and can provide distribution and abundance information about m6A methylation sites in transcripts. The method is suitable for identification and analysis of m6A methylation modification sites after virus infection, and can solve some problems brought to actual work due to special conditions that dengue virus typing is diversified and the virus does not have polyA tails.

The invention relates to the technical field of biology, in particular to a single cell methylation detection method based on enriched cast-off cells and application. The invention provides a single cell methylation detection method based on enriched cast-off cells. The single cell methylation detection method comprises the following steps: S1, separating, purifying and enriching cast-off cells to be analyzed; s2, carrying out DNA extraction on cast-off cells to be analyzed; s3, carrying out hydrosulfite conversion on the extracted DN A; s4, carrying out DNA multiple replacement amplification on the transformed DNA sample; and S5, carrying out methylation analysis on the amplified DNA. According to the method, DNA products are increased through a genome amplification technology, the DNA products can be increased to the level suitable for a conventional detection technology, and meanwhile non-specific loss in the capturing process is effectively avoided.

The present invention relates to the field of bioinformatics, and in particular to a methylome and metagenomics analysis method. The method comprises: obtaining data to be analyzed after preprocessing the whole genome sulfite sequencing data of the sample to be tested; and performing methylation analysis, bacterial and fungal microecological analysis, and phage analysis on the data to be analyzed. The present invention combines methylome and microbial metagenomics to analyze the whole genome sulfite sequencing data, and can effectively screen biological factors related to sepsis identification and related analysis. These biological factors can be used in sepsis identification, sepsis infection site prediction, and mortality analysis, and are of great value in the field of sepsis detection.

A method is provided for probabilistically assigning a tissue of origin to a nucleic acid in a sample, e.g., DNA in a cell-free fluid sample obtained from a human subject. A hydroxymethylation profile is generated for the sample DNA and then compared across a reference data set of hydroxymethylation profile vectors, where each hydroxymethylation profile vector identifies the hydroxymethylation profile at a specific reference locus, the tissue-specific gene associated with the reference locus, and the tissue with which the gene and reference locus are associated. A tissue of origin can be probabilistically assigned to the sample nucleic acid using the results of the comparison. Other methods of use are also provided.

To provide 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.SOLUTION: 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.SELECTED DRAWING: Figure 29A

The invention discloses a CpG methylation single cell whole genome analysis method and related equipment based on sequencing reads, and the method comprises the following steps: obtaining to-be-analyzed single cell data, and carrying out methylation data sequencing read comparison on the to-be-analyzed single cell to obtain a methylation site statistical result; screening a methylation specific interval based on a random sampling strategy and the methylation Hamming distance of the sequencing read; calculating a Hamming distance matrix of every two to-be-analyzed single cell CpG sites according to a methylation site statistical result and a methylation specific interval, and performing initial clustering based on the Hamming distance matrix; and obtaining cell DNA methylation heterogeneity measurement according to an initial clustering result, extracting a cell type or state specific DNA methylation mode panorama, calculating the similarity between a read segment and each cell type according to a specific methylation mode, and calculating the proportion of cells from a mixed sample sequencing result. According to the embodiment of the invention, the cell methylation analysis efficiency and sensitivity can be improved. The method can be widely applied to the technical field of gene analysis.

A method is provided for probabilistically assigning a tissue of origin to a nucleic acid in a sample, e.g., DNA in a cell-free fluid sample obtained from a human subject. A hydroxymethylation profile is generated for the sample DNA and then compared across a reference data set of hydroxymethylation profile vectors, where each hydroxymethylation profile vector identifies the hydroxymethylation profile at a specific reference locus, the tissue-specific gene associated with the reference locus, and the tissue with which the gene and reference locus are associated. A tissue of origin can be probabilistically assigned to the sample nucleic acid using the results of the comparison. Other methods of use are also provided.

The application discloses a sequencing read-based CpG methylation single-cell whole genome analysis method and related equipment, and the method comprises the following steps: obtaining single-cell data to be analyzed, performing methylation data sequencing read alignment on the single cells to be analyzed, and obtaining methylation site statistical results; screening a methylation-specific interval based on a random sampling strategy and a methylation Hamming distance of the sequencing read; calculating a Hamming distance matrix of CpG sites of each pair of single cells to be analyzed according to the methylation site statistical results and the methylation-specific interval, and performing initial clustering based on the Hamming distance matrix; obtaining a cell DNA methylation heterogeneity metric according to the initial clustering result, extracting a cell type or state-specific DNA methylation pattern panorama, calculating the similarity of a read and each cell type according to the specific methylation pattern, and calculating the proportion of cells from a mixed sample sequencing result. The embodiment of the application can improve the efficiency and sensitivity of cell methylation analysis. The application can be widely applied in the technical field of gene analysis.