27 results about "Dna cleavage" patented technology

The invention provides Cas12a mutants from Murraya bovis AAX08, and methods of using the Cas12a mutants. The Mb2Cas12a mutant comprises a single amino acid substitution, a plurality of amino acid substitutions, a structural domain exchange, or all of the above. Compared with a wild type moraxella bovine eye AAX08Cas12a enzyme, the mutants have enhanced DNA (Deoxyribose Nucleic Acid) lysis activity in plants.

Systems and kits are disclosed herein for genetic engineering (such as for DNA cleavage and gene-editing), which include catalytic nucleic acids and catalytic nucleic acid-assisting reagents. Methods of genetic engineering are also described, in which both catalytic nucleic acid-assisting reagents and catalytic nucleic acids are specific for a target site, thus, providing high-fidelity genetic engineering.

The present disclosure concerns polynucleotides and amino acids of Acidaminococcussp. Casl2a (Cpfl) and methods for their use for genome editing in eukaryotic cells. 20 24 20 14 49 05 M ar 2 02 4 A B S T R A C T 2 0 2 4 2 0 1 4 4 9 0 5 M a r 2 0 2 4

The present disclosure concerns polynucleotides and amino acids of Acidaminococcus sp. Cas12a (Cpf1) and methods for their use for genome editing in eukaryotic cells.

Disclosed herein are compositions and methods for treatment and prevention of low back pain. The compositions include vectors comprising nucleotide sequences encoding one or more CRISPR-Cas system guide RNAs and a RNA-directed nuclease. The methods include modulating expression of one or more genes in a cell using the compositions, introducing a CRISPR-Cas system into a cell comprising one or more vectors comprising the compositions, including site-specific DNA cleavage in a cell, and treating a subject having lower back pain, and lower back pain caused by degenerative disc disease using the compositions disclosed herein.

Provided are modified proteins that are functional in RNA-guided DNA cleavage. The proteins include modified IscBs protein that have a modification of the N-terminus or C-terminus, or both. The modifications include a truncation of a PLMP domain of the IscB protein, or a PLMP domain that is relocated to a position of the IscB protein that is not the N-terminus. The modified IscB protein can be provided as a component of a fusion protein. The modified IscB proteins are used with an ωRNA to modify a DNA substrate.

The present invention discloses an application of a chelating agent in inhibiting the catalytic activity of low-dimensional nanomaterials and DNA shearing. When the chelating agent is used to inhibit the catalytic activity of low-dimensional nanomaterials, the chelating agent is mixed with the low-dimensional nanomaterial, so that the electrons provided by the chelating agent interact with the dangling bonds of the defects of the low-dimensional nanomaterial to form covalent bonds, thereby inhibiting the catalytic activity of the low-dimensional nanomaterial. The application of the chelating agent in DNA shearing is to mix the chelating agent with the low-dimensional nanomaterial and then mix it with a DNA solution, so that the electrons provided by the chelating agent interact with the dangling bonds of the defects of the low-dimensional nanomaterial to form covalent bonds, thereby inhibiting the effect of the catalytic activity of the low-dimensional nanomaterial on DNA shearing. The present invention utilizes the chelating effect of the chelating agent on the defects of the low-dimensional nanomaterial to inhibit the catalytic activity of the defects of the low-dimensional nanomaterial, and utilizes this property to successfully inhibit the shearing effect of the low-dimensional nanomaterial on DNA, which is of great significance for the later development of related biological applications of nanozymes.

Here, the inventors developed a strategy to modify the chromatin status of HSPCs at the HBG promoters to reactivate HbF expression. In particular, the inventors used from 1 to 4 single guide RNA (sgRNA) molecules spanning from the -220 to the -20 region of the HBG promoters together with CRISPR-based epigenome editors, namely dCas9-CBPcore and Tet1-dCas9. The first editor is capable of inserting histone acetylation while the second editor is performing DNA demethylation. Both epigenetic marks are associated with active transcription and are present in fetal erythroid cells expressing HbF. This strategy allows the ex vivo modification of adult HSCs to recreate a fetal-like epigenetic context leading to HbF production. Moreover, it avoids the genotoxicity associated with classical genome editing tools relying on DNA cleavage. Thus, the present invention relates to a method of increasing the expression of gamma globin in a population of eukaryotic cells through epigenome editing.

Described herein are compositions and methods for treatment and prevention of low back pain. The compositions include vectors comprising nucleotide sequences encoding one or more CRISPR-Cas system guide RNAs and a RNA-directed nuclease. The methods include modulating expression of one or more genes in a cell using said compositions, introducing a CRISPR-Cas system into a cell comprising one or more vectors comprising said compositions, inducing site-specific DNA cleavage in a cell, and treating a subject having lower back pain, and lower back pain caused by degenerative disc disease using the compositions disclosed herein.

Isolation or in vitro assembly of the Cas9-crRNA complex of the Streptococcus thermophilus CRISPR3 / Cas system and use for cleavage of DNA bearing a nucleotide sequence complementary to the crRNA and a proto-spacer adjacent motif. Methods for site-specific modification of a target DNA molecule using an RNA-guided DNA endonuclease comprising at least one RNA sequence and at least one of an RuvC active site motif and an HNH active site motif; for conversion of Cas9 polypeptide into a nickase cleaving one strand of double-stranded DNA by inactivating one of the active sites (RuvC or HNH) in the polypeptide by at least one point mutation; for assembly of active polypeptide-polyribonucleotides complex in vivo or in vitro; and for re-programming a Cas9-crRNA complex specificity in vitro or using a cassette containing a single repeat-spacer-repeat unit.

The present invention relates to modified gRNA molecules, compositions, and methods for site-directed gene editing and genome modification, such as DNA cleavage and gene activation or repression. The modified guide RNA of the present invention has a modified secondary structure (e.g., a long upper stem and a modified hairpin structure) that specifically targets a target DNA sequence having reduced off-target activity.

The present disclosure relates to polynucleotides and amino acids of Acidaminococcus sp. Cas12a (Cpf1) and methods for their use for genome editing in eukaryotic cells.

The application relates to a preparation method and application of an electrochemical sensor for sensitive detection of lead ions based on a metal nanocomposite. 2+ The introduction of the target Pb 2+ triggers the S-DNA cleavage of the DNAzyme to cut off a single-strand DNA, and simultaneously initiates the DNA Walker effect to obtain a large amount of single-strand DNA. The supernatant after magnetic separation is added dropwise on an electrode modified with HP1, after incubation, the single-strand DNA serves as an initiation chain to initiate the CHA signal amplification reaction, so that more and more signal probes are connected to the electrode surface, and the electrical signal in the system is enhanced. The quantitative analysis of lead ions is realized by using the change of the electrical signal corresponding to the change of the methylene blue concentration on the electrode surface before and after the addition of Pb 2+ The prepared electrochemical sensor has high sensitivity, a wide detection range, good selectivity and excellent practical application capability.

The present disclosure concerns polynucleotides and amino acids of Acidaminococcus sp. Cas12a (Cpf1) and methods for their use for genome editing in eukaryotic cells.

The application discloses an ICD inducer Ru(II) complex with a light-driven DNA breaking ability, a preparation method and application thereof, relates to the field of antitumor drugs, and the structural formula of the Ru(II) complex is [Ru(dip)2(L)](PF6)2. The Ru(II) complex can enter a cell nucleus to break DNA and activate immunogenic death mediated by the pyroptosis and cGAS-STING pathways by using photodynamic therapy, so that the antitumor immune effect of the body is improved. In the in-vivo experiment, the immune cells in the tumor microenvironment of the mice in the vaccine group are good, and the antitumor immune effect of the three injections of the vaccine is better. The results of H&E staining show that the vaccine does not cause obvious pathological damage to the main organs of the mice, and the vaccine has good biological safety.

The invention relates to the technical field of tumor immunotherapy, and particularly discloses a screening method of a cancer cell mitochondrial DNA (deoxyribonucleic acid) coding new antigen, mitochondrial DNA with higher purity is obtained by means of microdissection, linear DNA splitting and the like, and nuclear metastasis mitochondrial fragment interference in mitochondrial sequencing is overcome. On the basis, mitochondrial amplicon sequencing is used for identifying the base sequence change caused by mutation from the DNA level, multiple types of tumor vaccines including polypeptide vaccines, RNA vaccines and dendritic vaccines can be prepared according to requirements, and even the tumor vaccines can be used for TCR-T therapy. According to the screening method of the mitochondrial DNA coding new antigen and the application of the mitochondrial DNA coding new antigen in preparation of the vaccine, the feasibility of the mitochondrial coding new antigen and the effectiveness of the mitochondrial coding new antigen as a tumor vaccine target are accurately and comprehensively confirmed, and a transcription translation process and a screening mechanism of DNA-RNA-polypeptide are systematically constructed; and more diversified, personalized, safe and effective treatment strategies can be selected and designed for the patient.

This invention provides a method for detecting microRNA using thermophilic TtAgo protein based on cofactor activation of its mesophilic DNA cleavage activity. This invention expands the applicable temperature range of TtAgo by using a cofactor that extends the function of the extremely thermophilic TtAgo protein and activates its mesophilic DNA cleavage activity, thereby establishing a one-step closed-tube isothermal multiplex analysis method for microRNA based on TtAgo that is not limited by high temperatures.

Described herein are CAS12A mutants from Lachnospiraceae bacterium and methods for use thereof. These mutants have enhanced DNA cleavage activities at non-canonical TTTT protospacer adjacent motifs (PAM) compared to the wild-type enzyme.

The invention discloses an ICD inducer Ru (II) complex with photodynamic DNA breaking capacity and a preparation method and application thereof, and relates to the field of antitumor drugs, the structural formula of the Ru (II) complex is [Ru (dip) 2 (L)] (PF6) 2, and the Ru (II) complex can enter cell nucleus to break DNA and activate pyroptosis and cGAS-STING pathway mediated immunogenic death when applied to photodynamic therapy, so that the antitumor immune effect of a body is improved. In an in-vivo experiment, the vaccine group has a good effect on immune cells of a tumor microenvironment in a mouse, and the three-needle vaccine has a better anti-tumor immune effect; hamp, Hamp; the E staining result shows that the vaccine has no obvious pathological injury on the main organs of the mouse, and the vaccine is proved to have better biological safety.

The present disclosure concerns polynucleotides and amino acids of Acidaminococcus sp. Cas12a (Cpf1) and methods for their use for genome editing in eukaryotic cells.

This disclosure relates to compositions (e.g., spike-in controls) for use in immunoprecipitation (IP) assays and improved methods for analyzing results therefrom. IP assays, such as a chromatin immunoprecipitation (ChIP) assay, a ChlPmentation assay, a ChlP-exo assay, an MNase-ChIP assay, a high resolution X-ChIP assay, an insertional ChIP (iChlP) assay, and an engineered DNA- binding molecule-mediated chromatin immunoprecipitation (enChIP) assay, and immunotethering assays such as a chromatin Immunocleavage (ChIC), a CUT&RUN assay, a CUT&Tag assay, and a chromatin integration labeling followed by sequencing (ChIL-seq) assay, are powerful techniques for understanding chromatin biology. While existing strategies for normalizing IP data use spike-in controls carrying specific histone modifications (e.g., post-translational modifications), similar strategies do not exist for spike-in controls utilizing chromatin-binding proteins. The compositions and methods described herein use chromatin-binding protein-based spike-in controls, which results in reduced technical variation and improved quantification of IP or DNA cleavage-based assays.

Described herein are CAS12A mutants from Lachnospiraceae bacterium and methods for use thereof. These mutants have enhanced DNA cleavage activities at non-canonical TTTT protospacer adjacent motifs (PAM) compared to the wild-type enzyme.

The present invention relates to an optimized technology for regulating target-specific gene expression based on an ultra-miniaturized OMEGA system, and a gene expression regulatory system that can enhance gene regulatory efficiency using TnpB is constructed. In the present invention, mutated TnpB with an inactivated DNA cleavage function, or engineered reRNA was developed, establishing the optimal conditions that can yield gene expression efficiency using original TnpB without any introduced mutations, and the vector for regulating gene expression of the present invention is small enough to be loaded in AAVs. Thus, it can be effectively used in a gene expression regulation technology.

The present disclosure relates to DNA-PK inhibitors of formula (I), (I), or a pharmaceutically acceptable salt, stereoisomer, solvate, prodrug, or tautomer thereof, methods of making the foregoing and compositions thereof, and methods of use of compounds of formula (I) in combination with DNA cleavage agents.

This invention pertains to mutant Cas9 nucleic acids and proteins for use in CRISPR / Cas endonuclease systems, and their methods of use. In particular, the invention pertains to an isolated mutant Cas9 protein, wherein the isolated mutant Cas9 protein is active in a CRISPR / Cas endonuclease system, wherein the CRISPR / Cas endonuclease system displays increased on-target editing activity relative to a wild-type CRISPR / Cas endonuclease system. The invention also includes isolated nucleic acids encoding mutant Cas9 proteins, ribonucleoprotein complexes and CRSPR / Cas endonuclease systems having mutant Cas9 proteins that display increased on-target editing activity relative to a wild-type CRISPR / Cas endonuclease system.

The present invention relates to modified gRNA molecules, compositions and methods for site-specific gene editing and genome modification, such as DNA cleavage and gene activation or inhibition. In some embodiments, the modified guide RNAs have modified secondary structures (e.g., long upper stems and modified hairpin structures) that specifically target a target DNA sequence with reduced off-target activity.

This invention specifically relates to the preparation of a CRISPR-immunoassay combined with small molecule detection probe, its related detection method, and its applications. The probe preparation mainly employs a chemical-biological conjugation method, using the bifunctional cross-linking agents 4-(N-maleimidemethyl)cyclohexane-1-carboxylic acid sulfonate succinimide sodium salt (Sulfo-SMCC) and N-succinimide-S-acetylthioacetate (SATA) to covalently conjugate dsDNA modified with -NH2 at the 5' end and a monoclonal antibody to prepare a dsDNA-antibody probe. Secondly, this detection method uses the dsDNA-antibody probe as a bridge to combine the immune system and the CRISPR / Cas12a system. It utilizes the antibody's specific recognition ability for small molecules, and leverages RPA nucleic acid amplification for signal amplification and the cis-cleavage activity and / or trans-single-strand DNA cleavage activity of CRISPR / Cas12a to generate a fluorescent signal, achieving the conversion of the immune response signal into a high-intensity fluorescent signal, thus establishing a CRISPR-immunoassay combined with small molecule detection system. The detection method has advantages such as simple operation, high specificity, extremely high sensitivity, and stable signal.