209 results about "Cytosine" patented technology

The invention relates to a metabolic marker for diagnosing diabetic secondary osteoporosis and application of the metabolic marker, and belongs to the technical field of biological medicine. The method comprises the following steps: firstly, screening out obviously different metabolites between osteoporosis (DOP) and osteoporosis-free groups (DM) in diabetic patients; and secondly, screening out significant difference metabolites between the primary osteoporosis patient and the healthy control group, and excluding the intersection of the two groups of difference metabolites to obtain metabolites which exclude the influence of the primary osteoporosis. Metabolic markers, namely cytosine nucleotide, ketoglutaric acid and triethylamine, are obtained through further screening, and when the three metabolic markers are independently used, AUCgt; the AUC during combined use is 0.99, so that the limitation that the sensitivity of the traditional bone mineral density detection on the osteoporosis recognition in the diabetic population is insufficient is effectively made up.

The invention provides a perovskite solar cell, a preparation method thereof and a photovoltaic module. The perovskite solar cell comprises a first electrode, a hole transport layer, a perovskite layer, an electron transport layer and a second electrode which are stacked in sequence. The material of the hole transport layer comprises a biomolecular material which is fixed on a self-assembly monomolecular layer material through an intermolecular force, and the biomolecular material comprises one or more of guanine, adenine, thymine, uracil, cytosine, hydroxymethylcytosine, xanthine, hypoxanthine, 6-mercaptopurine and corrin. According to the perovskite cell provided by the invention, the biomolecular material with a plurality of nitrogen heterocyclic structures and amino groups is added to generate multipoint hydrogen-bond interaction and pi-pi accumulation with the tail end of the SAM, so that the orderliness of molecular arrangement is improved, and the dipole direction and strength of the whole interface are regulated and controlled; and a coordination bond or hydrogen bond network is formed by the coordination compound and uncoordinated cations or anions in perovskite, so that interface defects are effectively passivated, and the thermal stability and the humid and hot life of a device are enhanced.

The invention relates to the field of gene engineering. In particular, the present invention relates to cytosine deaminases and their use in base editing. More specifically, the invention relates to a method for screening and identifying a deaminase, a base editing system based on a newly identified cytosine deaminase, a method for editing a target sequence in a genome of an organism (such as a plant) by using the base editing system, and a method for screening and identifying the target sequence. As well as genetically modified organisms (e.g., plants) and progeny thereof produced by the method.

The invention discloses a Cg12n-v4.6 nuclease. The amino acid sequence of the nuclease is as shown in SEQ ID No. 12. The invention also discloses a preparation method of the nuclease. The Cg12n-v4.6 nuclease mutant with increased activity and improved editing efficiency is developed, and the editing efficiency of the Cg12n-v4.6 nuclease mutant is up to 70% or above. Meanwhile, through engineering modification of corresponding sgRNA of Cas12n, the size of the sgRNA skeleton is reduced. An enhanced Cas12n gene editing tool, namely an eCg12n gene editing system, is developed by combining an engineered Cg12n-v4.6 nuclease mutant and a simplified sgRNA skeleton. Meanwhile, by fusing deaminase, a compact cytosine base editor and a compact adenine base editor are constructed, and the cytosine base editor is applied to edit and induce disease genes to generate early termination codons, so that safe knockout of the genes is realized. Compared with an original wild type editing system, the Cg12n-v4.6 nuclease mutant and a compact eCg12n gene editing system derived from the Cg12n-v4.6 nuclease mutant have the advantages that the editing efficiency is improved, the system size is further simplified, and the Cg12n-v4.6 nuclease mutant and the compact eCg12n gene editing system derived from the Cg12n-v4.6 nuclease mutant are more suitable for gene editing application.

The present invention belongs to the technical field of genetic engineering. Provided are a single-base editor, a deaminase used therein, and the use thereof. The technical problems to be solved are to identify a naturally occurring cytosine deaminase without sequence preference, construct a base editor, and improve the efficiency and scope of base editing. In order to solve the technical problems above, a cytosine base editor is provided. The cytosine base editor is a fusion protein, wherein the fusion protein is a protein containing a cytidine deaminase, a Cas protein and a uracil-DNA glycosylase inhibitor, and the cytidine deaminase is a protein having an amino acid sequence of positions 28-164 of SEQ ID NO. 2. Further provided is the use of the fusion protein above and a biomaterial related thereto in plant single-base editing. The single-base editor can improve the efficiency of cytosine base editing and accurately mediate the base mutation of a target, and is widely applicable in the cells of maize and even other plants.

Disclosed are methods of preparing a composition of non-naturally occurring DNA, comprising: (a) extracting DNA from a sample of a subject; (b) contacting the extracted DNA or its derivative with a panel of oligonucleotide probes designed to hybridize to a plurality of preselected genomic regions, thereby generating selected DNA, wherein at least 50% of the preselected genomic regions are CpG regions that each comprises at least 3 CpG sites and a CpG density of at least 0.02 CpG / bp, wherein the preselected genomic regions cover 10-500 Mb of sequence space in human genome, wherein the extracted DNA or its derivative or the selected DNA or its derivative is further treated with an agent or a combination of agents that discriminates between methylated and unmethylated cytosines; and (c) performing high-throughput sequencing on the selected DNA or its derivative and generating sequencing reads.

The invention discloses a high-efficiency wide-targeting-range C-to-G double-enzyme synergistic base editor and application thereof, and belongs to the technical field of gene editing. The editor is a fusion protein comprising a Cas protein having reduced or lost endonuclease activity, a cytosine deaminase (CDA), and a cytosine DNA glycosylase (CDG). In order to solve the problems that an existing C-to-G editor is low in efficiency and limited in targeting range, the C-to-G editor synergistically and efficiently generates a base removal (AP) site on a target DNA through the dual effects of CDA and CDG, so that C-to-G base transversion is promoted. According to the editor, the C-to-G editing efficiency is remarkably improved, an editing window can be effectively expanded or moved, the targeting flexibility is greatly enhanced, meanwhile, high genome specificity is kept, and the editor shows strong application potential in various organisms such as yeast and plants.

Provided are siRNA compositions comprising gemcitabine (GEM) substituted for cytosine moieties in siRNAs. Provided are pharmaceutical compositions containing these siRNA molecules, and methods of using these compositions to treat diseases, such as cancer.

The application discloses cytosine deaminase and related biological materials and applications thereof, and belongs to the technical field of proteins. The technical problem to be solved by the application is to mine natural sequence-unpreferred cytosine deaminase, and to improve base editing efficiency and range. The application provides a protein with amino acid sequence of SEQ ID NO. 2 at positions 15-143. The application also provides application of the above protein and related biological materials thereof in single base editing. The application uses macro-genome big data, performs comprehensive and systematic bioinformatics mining analysis on novel cytosine deaminase, and selects a protein L85 highly similar to known DddA protein structure but with low sequence similarity for function verification. The protein L85 has high catalytic activity as cytosine deaminase, and has no target sequence preference restriction, and can be widely applied to in-vitro and in-cell / tissue gene editing, and has good application potential and development value.

The invention relates to the technical field of dye separation, and discloses a loose nanofiltration membrane and a preparation method and application thereof, the loose nanofiltration membrane comprises a porous support membrane, and a polyesteramide separation layer formed on the porous support membrane through one-step interfacial polymerization of nucleoside monomers and multi-acyl chloride monomers. The nucleoside monomer is one or more than two of adenosine, guanosine and cytidine nucleoside. The nucleoside monomer with a long-chain structure, a branched-chain structure, a rigid twisted structure and a plurality of hydrophilic reaction sites is selected as an interfacial polymerization water-phase monomer, so that the loose nanofiltration membrane is endowed with excellent permeation and separation performance. The membrane has a high rejection rate on dye molecules such as Congo red and methyl blue, also has high water permeability and extremely high dye / inorganic salt separation selectivity, and shows good application potential in the aspects of dye production and purification and dye wastewater recovery.

The invention includes improved methods and compositions for reduction of a C5-C6 double bond of a cytosine. In particular, the improved methods and compositions for reduction of a C5-C6 double bond of a cytosine is via enzymatic means, not via chemical means. In particular, the disclosure is directed to methods of converting 5,6-dihydro-fC (fC) and / or 5,6-dihydro-caC to 5,6-dihydro-U (DHU). In particular, the disclosure is directed to methods of converting 5fC and / or 5caC to DHU. In addition, the disclosure is directed to methods for detection of epigenetic cytosine modification, particularly cytosine methylation, using ene reductases to reduce the C5-C6 double bond of cytosine.

The present invention relates to a modified mitochondrial comprising, on the outer membrane of the mitochondrial, an antibody that binds to a cancer cell-specific protein and a cytosine deaminase. It is proved that when the mitochondria and 5-FC are jointly used for treating a pancreatic cancer cell line, the cytotoxicity induced by 5-FC treatment is remarkably enhanced. The effect is proved as follows: 5-FC is converted into 5-FU by cytosine deaminase expressed on a mitochondrial outer membrane, so that pancreatic cancer cells are induced to generate apoptosis. Therefore, the modified mitochondria according to the present invention can be effectively used in anticancer therapy using 5-FC as an anticancer agent.

The present invention provides technology for inhibiting or suppressing proliferation of cells. Disclosed herein is a double-stranded RNA that has a first strand and a second strand which is complementary to the first strand, wherein the first strand has a main sequence which comprises 19-23 bases and in which a base at the 5'-end is guanine (G) or cytosine (C) and an additional sequence which comprises 2-4 bases and which is added to the 3'-end side of the main sequence. The main sequence is a portion of a base sequence that encodes semaphorin-3A, the portion including at least a portion of a base sequence that encodes a signal peptide region of the semaphorin-3A.

Provided are compositions, methods, and systems for DNA pyrimidine-base editing. In some embodiments, provided are engineered thymine-modifying polypeptides or engineered cytosine-modifying polypeptides comprising a variant of a uracil-DNA glycosylate (UNG). In some embodiments, provided are such engineered polypeptides and a DNA recognition domain, which are configured to target a nucleic acid for excision. Also provided herein are nucleic acid encoding the polypeptides described herein, additional components useful for editing, such as sgRNA, kits, medicines, composition, and method of use thereof.

The present invention is directed to improved materials and methods related to cytosine and adenosine base editing of genomic DNA.

Described herein are methods of modifying or editing a target nucleic acid such as methods that edit adenine to cytosine, thymine, or guanine. Compositions and systems for modifying or editing a target nucleic acid are also described. Methods, compositions and systems described herein may be used for generating allelic diversity.

A duchenne muscular dystrophy-related exonic splicing enhancer, sgRNA and gene editing tool can be applied in the preparation of drugs for treating duchenne muscular dystrophy. The gene editing tool designed on the basis of cytosine deaminase AID mutants and Cas9 mutants can perform site-specific modification on a mammalian genome by using an adeno-associated virus (AAV) as a vector. By optimizing an encoding nucleic acid sequence and an element composition structure of the editing tool, site-specific targeted modification of mammalian genetic material DNA can be efficiently achieved; and by performing targeted genetic manipulation on the nucleic acid sequence carrying disease mutations, a pathogenic mutation cannot be retained in a mature protein amino acid sequence or the pathogenic mutation cannot perform its function, so that the purpose of treating various gene mutation type genetic rare diseases is achieved, and the advantages of high efficiency, safety and stability are achieved.

本文描述了一种用于制备测序文库的方法。所述方法可包括：在连接缓冲液中使一种或多种衔接子连接至核酸分子（例如，DNA 或 RNA）；在所述连接缓冲液中使所述核酸分子与多个珠结合；从所述连接缓冲液中分离与所述核酸分子结合的所述多个珠；将所述多个珠悬浮在包含醇的溶液中；以及使用亚硫酸氢盐反应，使所述核酸分子中的未甲基化胞嘧啶转化为尿嘧啶，以生成转化的核酸分子。

Described herein is a base editing system for correcting mutations G3460A, G11778A, or T14484C in mitochondrial DNA of a patient with Leber hereditary optic neuropathy (LHON) to a normal genotype. Also, described herein is a method for correcting a mutation in the mitochondrial genes of a patient with LHON to a normal genotype using a base editor that recognizes specific sites in the mitochondrial genes of the patient with LHON and has an activity of specifically correcting the adenine base at position 3460 or 11778, or the cytosine base at position 14484, by using a fusion protein or a polynucleotide encoding such a fusion protein. The base editor or nucleotide described herein may correct DNA mutations specific to LHON in a cellular or extracellular in vitro environment. Thus, described herein is also the use of the substance in the prevention or treatment of LHON.

The invention discloses a construction method and application of a basic group editor of industrial bacteria of cephalosporium acremonium based on CRISPR-Cas9 (clustered regularly interspaced short palindromic repeats-associated protein 9), and belongs to the field of gene engineering. The cephalosporium acremonium industrial bacterium base editor provided by the invention can be used for specific mutation of a specific site of the DNA level of the cephalosporium acremonium industrial bacterium. The cytosine base editor CBE constructed by the invention is a plasmid comprising a fusion protein A and an sgRNA expression element. The base editor CBE can successfully mediate C.G-T.A, is suitable for the base editor of the high-yield CPC cephalosporium acremonium industrial bacteria, utilizes the protoplast, is high in conversion efficiency, can mediate specific site mutation in the cephalosporium acremonium industrial bacteria, and has application value.

Aspects of the present disclosure are directed to methods, compositions, and kits for detection and analysis of DNA and RNA cytosine methylation and / or RNA pseudouridylation. Certain aspects include methods, compositions and kits useful in bisulfite sequencing of methylated and / or pseudouridinylated nucleic acids, including methylated and / or pseudouridinylated nucleic acids from low-input samples such as cell-free DNA (cfDNA) and cell-free RNA, long fragment polynucleotides, and / or cfDNA and DNA from formalin-fixed paraffin-embedded (FFPE) samples. Also disclosed herein are methods, compositions and kits useful in bisulfite sequencing of methylated and / or pseudouridinylated nucleic acids with low nucleic acid damage, higher true positive results, and / or lower false negative results.

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.

This useful solution refers to a primer and probe set for multiplex methyl-specific qPCR (multiplex qMSP) to determine the methylation rate of cytosine in the promoter regions of two genes, FAM19A4 and MIR124-2, of the Vietnamese female genome.

The present application relates to a promoter pAtEF1 alpha for improving gene editing efficiency of dicotyledon plants and application thereof, and belongs to the field of plant genetic engineering.The pAtEF1 alpha promoter can be used for expression of adenine base editing protein, cytosine base editing protein, Cas nuclease and gRNA.The present application further provides an expression cassette group containing the pAtEF1 alpha promoter and a recombinant vector containing the expression cassette group.The pAtEF1 alpha promoter can be widely applied to the fields of gene editing and base editing of dicotyledon plants, and has wide application in genetic improvement of dicotyledon crops such as tomato and biosynthesis of high-value metabolites.

Provided herein are cytosine deaminase variants with enhanced preference for particular motifs, polynucleotides and vectors encoding the same, and methods of using the same.

The present invention relates to a DNA base editing composition including uracil DNA glycosylase (UDG), and a DNA base editing method using same. Specifically, the present invention relates to a DNA base editing composition including a DNA binding protein, cytosine deaminase, adenine deaminase, and UDG, or polynucleotides encoding the proteins, or a DNA base editing method using same. The present invention is useful for correcting bases in nuclear DNA or organellar DNA and particularly for correcting bases in DNA of organelles such as chloroplasts or mitochondria. In the present invention, the UDG is present independently without being fused to the DNA binding protein, the cytosine deaminase, and / or the adenine deaminase.

The invention relates to a UdgX-SSBE3 fusion protein, a method for capturing target DNA from a sample and application of the UdgX-SSBE3 fusion protein. The UdgX-SSBE3 fusion protein is formed by fusing a cytosine base editor BE3 and a uracil binding protein UdgX, and the method is a method for capturing DNA containing a specific target sequence (containing PAM and containing cytosine C at a specific position) from a sample at high flux. Comprising the following steps: 1) contacting a sample containing the specific target DNA with UdgX-SSBE3 protein to capture the target DNA, and 2) analyzing the target DNA through methods such as next-generation sequencing and the like. The invention also provides application of high-throughput capture and / or detection of DNA containing a specific target sequence from a sample through the base editing-uracil binding protein under mediation of the sgRNA library. The method provided by the invention can capture the target DNA from a complex nucleic acid sample with high flux, high sensitivity and high specificity, and has practical application value.

The invention discloses a lung nodule diagnosis method based on plasma DNA methylation characteristics and application, and the method comprises the following steps: S1, collecting peripheral blood of a subject in a special cfDNA preservation tube, carrying out centrifugal separation to obtain plasma, extracting cfDNA by adopting an optimized paramagnetic particle method or column extraction method, carrying out quality control on the cfDNA, then carrying out thorough hydrosulphite conversion treatment on the cfDNA, and carrying out purification treatment on the cfDNA to obtain the plasma; unmethylated cytosine is converted into uracil, and methylated cytosine is kept unchanged. The invention is derived from the selection of an early and stable epigenetic marker of DNA methylation, and replaces traditional dependent clinical parameters or mutation signals appearing at a late stage. According to the method, the problem of insufficient discrimination capability (AUClt; 0.60) caused by limited input information of an existing clinical prediction model (such as a MayoClinic model) is directly solved.

In some aspects and embodiments, the invention provides methods for making hematopoietic stem cells, including for HSCT. The method comprises providing a cell population comprising hemogenic endothelial (HE) or endothelial cells, and increasing activity or expression of DNA (cytosine-5-)-methyltransferase 3 beta (Dnmt3b) and / or GTPase IMAP Family Member 6 (Gimap6) in the HE and / or endothelial cells under conditions sufficient for stimulating formation of HSCs.

The present invention relates to a base correction system that corrects mitochondrial DNA mutations G3460A, G11778A, or T14484C, which are present in patients with Leber's hereditary optic neuropathy (LHON), to a normal genotype. Specifically, the present invention provides a base editor capable of correcting a mutation site in a mitochondrial gene of an LHON patient to a normal genotype. The present invention also provides a method for correcting a mitochondrial gene mutation using a fusion protein or a polynucleotide encoding such a fusion protein that recognizes a specific site in the mitochondrial gene of an LHON patient and specifically corrects the adenine base at position 3460, the adenine base at position 11778, or the cytosine base at position 14484. The base editor or polynucleotide according to the present invention can be used in cells or in an extracellular test tube environment to correct DNA mutations specifically expressed in LHON, and more preferably, can be used as a gene therapy agent to prevent or treat the disease. Thus, the present invention also provides a use of the substance for preventing or treating Leber's hereditary optic neuropathy.