38 results about "Bisulfite sequencing" patented technology

The invention relates to a sieving and checking method for cancers, comprising the following steps: (1), providing a test body to be checked; (2), checking the CpG sequence methylation state of at least one target gene in the genome DNA of the test body to be checked, wherein at least one of the target genes comprises SOX1, PAX1, LMX1A, NKX6-1, WT1 and ONECUT1; and (3), judging whether the test body has cancers or precancerous lesions or not according to the existence or inexistence of the methylation state of the target genes. The methylation state checking method comprises a methylation specific polymerase chain reaction, a quantitative methylation specific polymerase chain reaction, sulphite sequencing, micro-arrays, mass spectrograph analysis or denaturation high-efficiency liquid chromatography, and the like.

Systems and methods that analyze blood-based cancer diagnostic tests using multiple classes of molecules are described. The system uses machine learning (ML) to analyze multiple analytes, for example cell-free DNA, cell-free microRNA, and circulating proteins, from a biological sample. The system can use multiple assays, e.g., whole-genome sequencing, whole-genome bisulfite sequencing or EM-seq, small-RNA sequencing, and quantitative immunoassay. This can increase the sensitivity and specificity of diagnostics by exploiting independent information between signals. During operation, the system receives a biological sample, and separates a plurality of molecule classes from the sample. For a plurality of assays, the system identifies feature sets to input to a machine learning model. The system performs an assay on each molecule class and forms a feature vector from the measured values. The system inputs the feature vector into the machine learning model and obtains an output classification of whether the sample has a specified property.

The invention relates to a cancer screening method which comprises the following steps that: (1) providing a tested body; (2) testing the CpG sequence methylation state of at least one target gene in genome DNA of the tested body, and the target gene comprises PTPRR, ZNF582, PDE8B and DBC1; (3) judging whether the tested body has cancer or lesions before cancer according to that whether the methylation state of the at least one target gene exists; and the methylation state detection method is methylation-specific PCR, MSP, quantitative methylation-specific PCR, QMSP, bisulfite sequencing (BS), microarrays, massspectrometer, denaturing high-performance liquidchromatography (DHPLC) and the like.

The invention relates to a library construction method, a sequencing method and a methylation detection method for sequencing 5-methylcytosine (5mC) modified bisulfite on RNA molecule. By designing and using triplet base random hexamers which contain ACT only, RNA fragment reverse transcription, PCR amplification efficiency and 5mC site detection efficiency after bisulfite treatment can be significantly improved, which is conducive to high-throughput sequencing of the 5mC site.

The invention provides a method for simultaneously detecting a methylation level, genome variation and an insert fragment by utilizing a methylation non-bisulfite sequencing technology. The genome variation comprises gene mutation, copy number variation and structural variation. The invention also provides a device and equipment for implementing the method, and a corresponding computer readable medium. According to the invention, for methylated non-bisulfite sequencing data, one-step detection and analysis from offline data to multiple dimensions of methylation level, genome variation and insert fragments are realized, the method is suitable for whole genome and targeted capture data types of methylated non-bisulfite, and analysis of a single cancer sample and paired samples (cancer samples containing control samples) can be carried out.

The invention relates to a kit for morbidity-related tumor suppressor gene epigenetic mutation detection of gastrointestinal neoplasms and application thereof. The kit mainly comprises 25 pairs of gastrointestinal neoplasm morbidity-related tumor suppressor gene promotor region CpG-island amplification and sequencing primers of 9 kinds and a polymerase chain reaction (PCR) amplification reagent. Through a PCR amplification product obtained through the kit, a gene CpG-island methylation state can be analyzed through the methods of methylation specific PCR (MSP), combined bisulphite-restrictionanalysis (COBRA) or bisulfite sequencing PCR (BSP) and the like. The kit has the efficient effect of detecting CpG-island methylation and is used for the morbidity-related tumor suppressor gene epigenetic mutation detection of the gastrointestinal neoplasms.

The present invention relates to a cancer biomarker molecule, a screening test kit and a detection method thereof. According to the present invention, analysis of methylating areas of specimen target genes PAX1, ZNF582, SOX1 and NKX6-1 are adopted, plural of segments of target gene-specific oligonucleotide primers or probes are designed, and the oligonucleotide probes are adopted to detect presence or absence of methylation of the target genes so as to further determine possibility of cancer occurrence; and the methylation state detection method comprises methylation-specific PCR (MSP), quantitative methylation-specific PCR (QMSP), bisulfite sequencing (BS), microarrays, mass spectrometer, denaturing high-performance liquid chromatography (DHPLC), pyrosequencing or Enzyme-linked immunosorbent assay (ELISA) and the like.

The invention belongs to the technical field of biomedicine. The abnormal methylation of genes participates in the occurrence and the development of tumors. The invention aims at overcoming the defects of the methylation sensitivity restriction enzyme method, the bisulfite sequencing method and the methylation specificity PCR (Polymerase Chain Reaction) method and providing an LRRC55 gene methylation quantitative detection method with easy and simple operation and high sensitivity. The method comprises the following steps of: taking a pancreatic cancer tissue, extracting DNA, preparing a standard curve sample of ACTB (Actin, Beta), and preparing a standard curve sample of LRRC55 by using a complete methylation template; extracting the DNA of a tissue to be tested, performing chemical modification, designing a methylation primer and probe aiming at the CpG island of a promoter region of the human LRRC55 gene, designing a BSP primer and probe aiming at the CPG island of a promoter region of human reference gene ACTB, performing real-time quantitative PCR on the DNA treated by sulfite by using the Tagman-MGB real-time quantitative PCR method, and calculating the quantitative value of the methylation degree of the LRRC55 gene. The method is rapid and accurate and is high in sensitivity.

The invention relates to a unicell simplified representative bisulfite sequencing library construction method which comprises the steps of unicell pyrolysis, MspI enzyme digestion, tail end repair, 3'- terminal A addition, methylated linker addition, bisulfite treatment and PCR (polymerase chain reaction) amplification. The invention also relates to a unicell simplified representative bisulfite sequencing library construction kit, and a detection method and detection kit thereof, belonging to the field of gene sequencing. By using the unicells as the research object, the unicell simplified representative bisulfite sequencing library construction method for sequencing can be used for researching cell heterogeneity on the unicell level.

The invention which belongs to the technical field of biological detection relates to a method for detecting MP of sheep and provides the methylation state of CpG islands in an MBL (mannan-binding lectin) gene promoter region related with the MP of sheep and an application thereof. The invention also provides a method for detecting the MP of sheep and concretely relates to the methylation detection of the MBL promoter region and the artificial infection verification. The methylation state detection of the MBL gene promoter region of sheep is carried out through selecting a bisulfite sequencing process, the methylation state and the frequency of each CpG locus in a given region can be accurately detected, three substantially differential CpG methylation loci, CpG3, CpG4 and CpG5, related with the MBL level are found, a case that the MBL level after toxic elimination is lower than the MBL level before the toxic elimination is promoted, and the MBL gene promoter region methylation may be a more substantial characteristic of the MBL level reduction of serum. The method allows the MP infection of sheep and susceptible individuals to be sensitively and accurately detected, and provides complete information and scientific bases for the sheep MBL gene promoter region methylation and the MP infection research of sheep.

The invention provides a method for detecting additive and dominant genetic effects of DNA methylation sites on quantitative traits and an application thereof, and belongs to the field of plant molecular breeding. The method comprises the following steps: 1) collecting population samples of germplasm resources, detecting phenotypic traits, and extracting genomic DNA from the samples; 2) constructing a bisulfite sequencing library by the genomic DNA of the samples, and sequencing; 3) identifying the DNA methylation sites from DNA methylation sequencing data, and genotyping; and 4) carrying outcorrelation analysis of the obtained genotyping data of the DNA methylation sites and the sample phenotypic trait data by a mixed linear model in Tassel 5.0 software package, and screening significantly correlated DNA methylation sites and analyzing the additive and dominant genetic effects. The method can provide new technical guidance for gene marker assisted breeding, and has important theoretical and breeding value.

The invention discloses a method for performing DedscRRBS-PGS analysis on an embryo culture solution. The invention provides a method for performing simplifying representative bisulfite sequencing andchromosomal aneuploidy status analysis on the embryo culture solution, and the method comprises the steps of 1) constructing a DNA library by using a blastocyst culture solution as a sample, and in the process of constructing the DNA library, using two restriction enzymes to enrich CpG sites by digesting genomic DNA of the blastocyst culture solution; 2) conducting CT conversion on the DNA library with bisulfite to obtain a conversion product; 3) using the conversion product for preparing a methylation sequencing library, following by high-throughput sequencing, and analyzing DNA methylationand a chromosomal aneuploidy status based on a sequencing result. The blastocyst culture solution is used as a raw material, a double enzyme digestion simplifying representative bisulfite sequencing technology is adopted, and the development potential of an embryo is comprehensively evaluated from multiple angles.

The invention provides an FOXP3 (forkhead/winged helix transcription factor 3) gene methylation detection method based on bisulfite sequencing. The method comprises the following steps: 4 mL of human venous blood is collected by a coagulation-promoting tube, solidified at room temperature for 30 min and centrifuged at 1,000 g*15 min; genomic DNA of a blood clot is extracted; integrity, concentration and purity of the genomic DNA are determined primarily; the genomic DNA is modified with a methylation kit; methylation PCR amplification is performed, a PCR product is identified with 2% agarose gel electrophoresis, and a gel imaging system performs observation and shooting; 18 mu L of the PCR product is purified and recovered by a DNA purifying and recovering kit, 2 mu L of the purified and recovered PCR product is transformed into Trans1-T1 competent cells through vector linking, blue-white clone plaque selection is performed, and positive clone is identified; a kanamycin monoclonal antibody LB liquid medium with the concentration of 50 mu g/mL is inoculated with thalli in positive clone plaques, the thalli are subjected to shake culture at the constant temperature of 37 DEG C and the rate of 200 rpm/min for 8-12 h, and samples turning to be turbid are taken for sequencing. Methylation degrees of 5 CpG sites of a specific sequence of a promoter region of an FOXP3 gene can be detected successfully with the method.

The present disclosure provides methods and systems for detecting or inferring levels of Copy Number Variants (CNVs) in cell-free nucleic acid samples to detect or assess cancer and prenatal diseases. Cell-free nucleic acid methylation sequencing data may be utilized to distinguish tumor-derived or fetal-derived sequencing reads from normal cfDNA sequencing reads. Each cell-free nucleic acid sequencing read (e.g., containing tumor or fetal methylation markers) may be classified as corresponding to a tumor/fetal-derived or a normal-plasma cell-free nucleic acid, based on the methylation cfDNA sequencing data (e.g., obtained using Bisulfite sequencing or bisulfite-free sequencing methods) and tumor/fetal methylation markers. Next, a profile of the tumor/fetal-derived sequencing read counts may be constructed and then normalized. The CNV status (e.g., gain or loss) of each genomic region may be inferred, and a diagnosis or prognosis can be made based on a subject's inferred CNV profile.