130 results about "Bisulfite" patented technology

The invention discloses a high-activity TET dioxygenase mutant as well as a construction method and application thereof. The TET dioxygenase mutant is obtained from an amino acid sequence of wild type TET dioxygenase as shown in SEQ ID NO.1 according to any one of the following mutation modes: a, the 150th proline is mutated into alanine; and b, proline at the 150th site is mutated into alanine, and isoleucine at the 235th site is mutated into methionine. The TET dioxygenase mutant provided by the invention can be stabilized in an escherichia coli system, the purification effect is good, the preparation difficulty is reduced, and the activity is remarkably improved compared with that of a wild type. The methylation gene transformation effect is far better than that of a traditional bisulfite transformation method, and the method has a good application prospect.

The invention discloses a tetrahydrocarbazole enhanced fluorescent probe for detecting hydrosulfite as well as a preparation method and application of the tetrahydrocarbazole enhanced fluorescent probe. The fluorescent probe is a 2-(2-(5-(2, 2-dimethyl-2, 3, 4, 9-tetrahydro-1H-1, 3-methyl carbazole-6-yl) furan-2-yl) vinyl)-3-ethyl benzothiazole-3-onium iodised salt compound, namely, TC-FV-BT, and the fluorescent probe can be applied to the field of fluorescence detection. The probe TC-FV-BT can be used for specifically recognizing the hydrosulfite. Under sunlight, after bisulfite is added into a DMF / H2O (2: 8, v / v) solution of the probe TC-FV-BT, the color of the solution is changed from dark pink to white, under irradiation of ultraviolet light with the wavelength of 365 nm, the solution emits orange fluorescence at the emission wavelength of 605 nm, the fluorescence color of the solution is changed from colorless to orange, the response time is 4 min, the detection limit of the bisulfite reaches 77.27 nM, and the detection limit of the bisulfite reaches 77.27 nM. The fluorescent probe shows good selectivity and anti-interference performance, and has a good application prospect in the fields of environmental analysis, food and the like as a fluorescent probe for detecting hydrosulfite.

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 belongs to the technical field of microneedle preparations, and particularly discloses a soluble microneedle for treating skin acnes and a preparation method and application thereof.The soluble microneedle comprises a hyaluronic acid compound, iron ions and bisulfite ions, the hyaluronic acid compound comprises aldehyde hyaluronic acid and binary polyphenol, and the iron ions and the bisulfite ions are combined to form the soluble microneedle. The hydroformylated hyaluronic acid is hyaluronic acid subjected to ring opening oxidation. According to the preparation method, hyaluronic acid is subjected to ring-opening oxidation to obtain hyaluronic acid carrying aldehyde groups, dopamine hydrochloride is chemically modified, iron ions are used as bridges to connect bisulfite ions, an antibacterial polymer is developed to serve as a matrix to be used for preparing the soluble microneedle, the soluble microneedle is punctured into the skin in a pressing mode, and the antibacterial effect is achieved. The substrate for manufacturing the microneedle is soluble, so that sharp waste is not generated, and the risk of secondary infection and the cost of biological waste treatment are reduced.

The invention provides a reverse osmosis reducing agent and a preparation method and application thereof, and belongs to the technical field of reverse osmosis water treatment.The reverse osmosis reducing agent comprises a first reducing agent, a second reducing agent and a solvent; the first reducing agent is thiosulfate, and the second reducing agent is sulfite and / or bisulfite; the mass ratio of the first reducing agent to the second reducing agent is (1: 18)-(10: 1). The reverse osmosis reducing agent can be applied to reverse osmosis water treatment, the reverse osmosis reducing agent is added into reverse osmosis inlet water to be treated, and the reverse osmosis inlet water subjected to reduction treatment can be obtained after full reaction. The first reducing agent and the second reducing agent are compounded according to the specific mass ratio, so that the compounded reverse osmosis reducing agent can effectively inhibit the occurrence of advanced oxidation chain reaction while keeping high reducibility, and effective reduction of reverse osmosis inlet water can be realized by using a small amount of the reverse osmosis reducing agent.

The present application relates to a method and system for inferring the age of an individual from semen or a semen stain, and a methylation detection kit, which comprises the following steps: firstly, extracting the genomic DNA of the semen or semen stain of the individual; secondly, performing bisulfite treatment on the genomic DNA; thirdly, selecting a plurality of sites in the CpG sites in the genomic DNA and amplifying the selected CpG sites to obtain the methylation rate of the selected CpG sites; and finally, performing regression analysis on the methylation rate of the selected CpG sites and the age of the individual to construct a regression model for inferring the age of an unknown individual from semen or a semen stain. The average absolute error of the present application reaches 1.68 to 4.44 years.

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.

The invention discloses a method for preparing a high-purity pentoxifylline intermediate, wherein the prepared 6-bromo-2-hexanone crude product is reacted with an alkali metal bisulfite to generate a novel 6-halogenated-2-hydroxyhexane-2-sulfonic acid alkali metal salt, and then the 6-bromo-2-hexanone pure product is generated under the action of a hydrohalic acid or a base, without the need for rectification and purification, and the post-processing operation is simple, in line with the trend of green chemical industry, and has a good industrial production application prospect. At the same time, the present invention can effectively avoid the dimer impurities generated in the high-temperature distillation process, and the yield and purity are high, which provides favorable guarantees for the safety and effectiveness of preparing the finished drug pentoxifylline.

A method of performing bisulfite sequencing on ultrashort single-stranded cell-free DNA (uscfDNA) and mononucleosomal cell-free DNA (mncfDNA) is described as well as methods of using the methylation profile of uscfDNA and / or mncfDNA for detecting biomarkers and diagnosing diseases and disorders.

The application discloses a free DNA methylation and fragmentomics combined analysis method based on targeted CpG bisulfite sequencing, and belongs to the technical field of liquid biopsy and epigenetic detection. The application extracts cfDNA from blood plasma, carries out high-throughput sequencing after library construction by a methylation adapter, bisulfite conversion, and biotin probe targeted capture of a CpG enrichment region, and simultaneously realizes single-base resolution methylation accurate quantification and FRAGMA fragmentomics analysis by using the same sequencing data, so that the accuracy of methylation quantification results is inferred, and the optimal detection region is iteratively selected; multi-dimensional characteristics such as a methylation ratio, an 11nt cutting map, a CGN / NCG motif ratio and a fragment length distribution are fused to construct a machine learning / deep learning model to output a unified disease risk score. The application realizes the bimodal integration of methylation chemical signals and fragmentomics structural signals in a targeted sequencing system for the first time, and has the advantages of low cost, high sensitivity and strong clinical adaptability.

The invention belongs to the technical field of gene methylation detection, and particularly relates to a cervical cancer related gene methylation detection kit and system, which comprises primers and probes for detecting APC, CALCA, DAPK and ESR1 gene methylation, primers and probes for monitoring internal control genes, a PCR reaction liquid, a positive quality control product and a negative quality control product, the use method comprises the following steps: 1) performing nucleic acid extraction on a sample; (2) converting the extracted genome DNA by bisulfite; (3) premixing and subpackaging the gene methylation detection reagent to obtain a PCR reaction system; (4) adding the DNA obtained in the step (2) into the PCR reaction system in the step (3), uniformly mixing, and carrying out PCR amplification; 5, gene methylation interpretation is carried out after the reaction is ended, the method has the advantages of being high in sensitivity, low in cost, easy and convenient to operate, short in detection period, visual in interpretation and the like, and methylation of the cervical cancer related genes can be rapidly and accurately detected.

The application discloses a cervical cancer methylation detection kit and application thereof, the kit takes PAX5 gene, PAX1 gene, MICOS10-NBL1 gene and MICOS10 gene as detection targets; the kit mainly comprises specific primer probe dry powder combination, PCR reaction reagent dry powder, positive control product dry powder, negative control product dry powder and microdroplet generation oil. The sample is detected by bisulfite conversion, and the kit is combined with microdroplet digital PCR detection technology to realize accurate detection of the methylation level of the target gene. The detection verification result shows that the sensitivity of the kit for cervical cancer detection reaches 96.9%, the specificity reaches 95.7%, the kit has the advantages of accurate detection, simple operation and strong stability, can be widely applied to early screening, clinical auxiliary diagnosis and physical examination of healthy people of cervical cancer, and provides reliable technical support for early discovery and early treatment of cervical cancer.

The invention relates to the technical field of colorectal cancer auxiliary detection, and particularly discloses a blood methylation site combination and a detection kit for early diagnosis of colorectal cancer, and the method comprises the following steps: providing a blood sample of a person to be detected for obtaining cfDNA; the cfDNA is subjected to bisulfite treatment, and the transformed DNA is obtained; amplifying the transformed DNA by using a multiplex fluorescence PCR (Polymerase Chain Reaction) method to obtain fluorescence signal data of each gene; calculating a Ct value of each gene based on the fluorescence signal data, comparing the Ct value with a preset Cut-off value, and judging whether the colorectal cancer exists or not; according to the present invention, the invasive operation of the colonoscope can be replaced, the screening can be completed through the blood sample, the patient compliance can be significantly improved, the method is suitable for the large-scale early screening, the trace tumor signal in the blood can be efficiently captured based on the methylation marker highly related to the colorectal cancer, and the accurate early canceration identification can be achieved.

The methods, compositions, and kits of the disclosure provide a novel approach for a whole genome, unbiased DNA analysis method that can be performed on limited amounts of DNA can be used to analyze DNA to determine its modification status. Aspects of the disclosure relate to a method for amplifying bisulfite-treated deoxyribonucleic acid (DNA) molecules comprising: (a) ligating an adaptor to the DNA molecules, wherein the adaptor comprises a RNA polymerase promoter comprising bisulfite-protected cytosines; (b) treating the ligated DNA molecules with bisulfite; (c) hybridizing the bisulfite-treated DNA molecules with a primer; (d) extending the hybridized primer to make double stranded DNA; and (e) in vitro transcribing the double-stranded DNA to make RNA.

The invention relates to a continuous preparation method of pentaerythritol bisulfite, which comprises the following steps: respectively and continuously introducing pentaerythritol emulsion and thionyl chloride into a continuous flow oscillation stirring reactor for continuous flow reaction, controlling the reaction temperature to be 40-90 DEG C, controlling the temperature to be 40-90 DEG C, and controlling the reaction time to be 1-3 hours; the stirring rotating speed is 200 to 800 r / min, the reaction pressure is 0.3 to 0.8 MPa, and the reaction time is 30 to 120 minutes. By using the continuous flow oscillation stirring reactor and specific reaction conditions, the continuous preparation is realized, and the pentaerythritol conversion rate, the product yield and purity, the reaction efficiency and the post-treatment simplicity are considered at the same time. The method has the remarkable advantages of simple process, low reaction operation difficulty, high reaction efficiency and the like, and can meet the comprehensive requirements of large-scale continuous industrial production on efficiency, cost and product quality.

The invention discloses an LAMP (loop-mediated isothermal amplification)-based rapid detection primer composition and kit for non-muscle invasive bladder cancer and application of the LAMP-based rapid detection primer composition and kit. The primer composition comprises a primer group for targeted amplification of a VIM gene and a TMEFF2 gene. According to the method, rapid nucleic acid extraction is achieved through a one-step method reagent, enzymatic methylation conversion is combined to replace a traditional bisulfite method, and nucleic acid damage and detection complexity can be remarkably reduced. A double-target detection system is constructed on the basis of VIM and TMEFF2 genes, and visual rapid detection of the non-muscle invasive bladder cancer is realized in combination with an LAMP detection technology. The whole detection process does not need a centrifugal machine or a PCR instrument, extraction, conversion and detection can be completed only through a metal bath or a water bath kettle, operation is easy and convenient, and cost is low. The kit provided by the invention is suitable for household self-test, reduces the detection burden of patients, and has wide clinical and household application prospects.

Methods and systems provided herein address current limitations of bisulfite-based methylation sequencing by improving the quality and accuracy of nucleic acid methylation sequencing and uses thereof for detection of disease. Methods that include minimally-destructive conversion methods for methylation sequencing as well as specialized UMI adapters provide for improved quality of sequencing libraries and sequencing information. Greater accuracy and more complete methylation-state information permits higher quality feature generation for use in machine learning models and classifier generation.

The invention relates to the technical field of molecular biological detection, in particular to a detection method for DNA methylation of a miR-34a gene promoter region and application. According to the method, the bisulfite reaction system is treated by two tubes, and the strategy of adjusting the reaction time according to the DNA dosage is combined, so that the conversion efficiency of the unmethylated cytosine is improved to 90% or above, and is obviously superior to that of the existing single-tube reaction.

This invention discloses a class of novel small-molecule fluorescent compounds based on thiazolyl-2-pyridone rings and their applications in analytical detection and diagnostics. The small-molecule fluorescent compounds of this invention have novel structures, readily available starting materials, mild synthesis conditions, and high yields. Some of the obtained compounds are responsive to amines, hydrazine, hypochlorite, and bisulfite, and are expected to serve as detection probes for these compounds. Other compounds exhibit long fluorescence emission and large Stokes shifts, effectively avoiding background spectral interference, generating Type I reactive oxygen species, and possessing near-infrared emission characteristics, showing potential as Type I photodynamic therapy photosensitizers.

Methods and systems provided herein address current limitations of bisulfite-based methylation sequencing by improving the quality and accuracy of nucleic acid methylation sequencing and uses thereof for detection of disease. Methods that include minimally-destructive conversion methods for methylation sequencing as well as specialized UMI adapters provide for improved quality of sequencing libraries and sequencing information. Greater accuracy and more complete methylation-state information permits higher quality feature generation for use in machine learning models and classifier generation.

A method for the continuous manufacture of taurine, comprising at least one of (a) – (c): (a) reacting an ethylene oxide and an alkali metal bisulfite to obtain a first intermediate product stream comprising an alkali metal isethionate; (b) reacting the alkali metal isethionate from the first intermediate product stream with ammonia to obtain a second intermediate product stream comprising an alkali metal taurate; and (c) converting the alkali metal taurate from the second intermediate product stream to obtain taurine and at least one by-product, wherein the by- product is suitable for reacting sulfur dioxide to alkali metal bisulfite, and separating the at least one by-product from the taurine to obtain at least one by-product stream comprising the at least one by-product and a product stream comprising the taurine; and an apparatus for the continuous manufacture of taurine.

The invention discloses a QPCR (Quantitative Polymerase Chain Reaction)-based DNA (Deoxyribose Nucleic Acid) methylation rate detection method, which comprises the following steps of: treating a DNA sample to be detected by bisulfite, dividing the DNA sample into a first sample and a second sample which are parallel to each other, and performing substrate amplification on a methylated DNA template and a non-methylated DNA template in the samples through PCR amplification by adopting the same amplification primer under the same condition; then respectively depleting dATP in a sample A reaction system and dGTP in a sample B reaction system through a template and a primer with thymine deoxyribonucleotide repetitive sequences or a template and a primer with cytosine deoxyribonucleotide repetitive sequences, and controlling independent detection of methylation and non-methylation templates through a d3TP system lacking dATP or dGTP; and finally, directly analyzing the fluorescence signal of the methylation template and the fluorescence signal of the non-methylation template, and calculating the methylation rate of the DNA sample to be detected through a methylation rate calculation function.

The present invention provides a method for improving the stability of a bird's nest acid aqueous solution, the method for improving the stability of a bird's nest acid aqueous solution comprising the step of adding a stabilizer to the bird's nest acid aqueous solution, wherein the stabilizer is one or more of a sulfite and a bisulfite. The present invention effectively suppresses the loss and discoloration of bird's nest acid by adding a sulfite and / or a bisulfite to the bird's nest acid aqueous solution, especially under the condition of low pH value and high bird's nest acid content, the effect is significantly better than the prior art, and provides technical support for adding bird's nest acid to cosmetics, broadening the application scope of bird's nest acid.

Methods of determining methylation of DNA are provided that include filtering a biological sample comprising a nucleic acid with a first matrix material to purify the DNA; eluting and denaturing the DNA to produce eluted denatured DNA; heating the DNA in the presence of bisulfite ions to produce deaminated nucleic acid; optionally contacting said deaminated nucleic acid with a second matrix material; desulphonating and eluting the bound deaminated nucleic acid; and performing methylation specific PCR, nucleic acid sequencing, and / or high resolution melting analysis (HRM) on said bisulfite-converted nucleic acid to determine the methylation of said nucleic acid, wherein multiple steps may be performed in a single reaction cartridge.

The invention belongs to the technical field of ion detection, and particularly relates to a preparation method of a long-wavelength emission fluorescent probe for rapidly detecting bisulfite, which comprises the following steps: dissolving isophorone and malononitrile in ethanol to obtain a product 1; 3-bromoquinoline and 5-formyl-2-thiopheneboronic acid are dissolved in a mixed solvent of dry methanol and dry toluene, and a product 2 is obtained; dissolving the product 2 in dichloromethane, and adding methyl trifluoromethanesulfonate to obtain a product 3; and dissolving the product 1 and the product 3 in ethanol, and adding piperidine to obtain the required fluorescent probe LSQ. Dissolving the obtained fluorescent probe LSQ in DMSO to prepare mother liquor A; a target product is obtained. The fluorescent probe provided by the invention has the advantages of high-selectivity rapid response to bisulfite, long-wavelength emission, no organic solvent detection, simple synthesis route and simple and convenient detection equipment and method, and can be used for qualitative and quantitative detection of bisulfite in food and biological systems.

The present application belongs to the field of medicine and chemical industry, and relates to a method for purifying losartan. In the method, bisulfite or sulfite is used to remove color impurities of losartan, and the method has the advantages of mild treatment condition, short operation period, high decolorization efficiency and suitability for industrial production.

Provided are a method for preparing Bisulfite DNA and a use thereof. In the method, the process is simplified and optimized, and the same reagent is used as a reagent having both lysis and transformation functions, thereby avoiding the conversion of multiple systems, and reducing the steps of washing, elution and the like of a DNA sample.

Disclosed are methods and systems for detecting methylation of the promoter of O-6-methylguanine-DNA methyltransferase gene (MGMT). In particular, the methods and systems may be utilized to detect methylation in the MGMT promoter in a DNA sample from a glioblastoma and optionally in order to predict whether a subject having the glioblastoma will respond to treatment with an alkylating agent. The methods and systems typically include a step of deep-sequencing the DNA sample after the DNA sample has been treated with a reagent that converts unmethylated cytosine to uracil such as a bisulfite reagent.