10 results about "Inverted Repeat Sequences" patented technology

This disclosure generally relates to super minimal transposon inverted repeat sequence (ITR) polynucleotides, compositions comprising the polynucleotides and methods of using compositions comprising the polynucleotides for the ex vivo and in vivo delivery of nucleic acids to cells, in particular, in vivo delivery of therapeutic genes to treat genetic disorders or diseases.

A method for improving the replication of covalent closed circular plasmids is provided. [Solution] The method comprises the step of preparing a covalent closed circular plasmid having a Pol I-dependent origin of replication and an insert containing a structured DNA sequence selected from the group consisting of a reverse repeat sequence, a directional repeat sequence, a homopolymer repeat sequence, a eukaryotic origin of replication, or a eukaryotic promoter enhancer sequence, wherein the structured DNA sequence is located at a distance of less than 1000 bp from the Pol I-dependent origin of replication in the direction of replication. The method also comprises the step of modifying the covalent closed circular recombinant molecule so that the Pol I-dependent origin of replication is replaced with a Pol III-dependent origin of replication, thereby improving the replication of the covalent closed circular plasmid with the resulting Pol III-dependent origin of replication. A covalent closed circular recombinant DNA molecule without antibiotic markers is also provided.

To provide nucleic acids (including AAV expression cassettes), AAV vectors, and compositions for use in methods of treating and / or delaying the onset of diseases associated with mutations in mecp2 genes, such as Rett Syndrome.SOLUTION: Also provided herein are methods for treating a brain-derived neurotrophic factor (BDNF) - associated disease and / or delaying the onset of a brain-derived neurotrophic factor (BDNF) - associated disease. The present disclosure provides a nucleic acid comprising an adeno-associated virus (AAV) expression cassette, wherein the AAV expression cassette comprises, in a 5' to 3' direction, a 5' inverted terminal repeat (ITR), a synthetic activity-dependent promoter, a Rett syndrome-associated gene, and a 3' ITR.SELECTED DRAWING: None

The invention belongs to the technical field of biological medicine, and particularly relates to the field of gene therapy and pain therapy. The invention firstly provides a new application of the ASIC3 gene in anti-inflammation. The invention further provides a sgRNA and AAV-CRISPRi virus system for targeted inhibition of the ASIC3 gene, the AAV-CRISPRi virus system is a CRISPRi system based on AAV delivery, and the core functional structure sequence of the AAV-CRISPRi virus system is a 5 '-ITR inverted repeat sequence, an EF1 alpha core promoter, a ZIM3 transcription inhibition structural domain, a linker, a sadCas9-NLS nuclear localization signal, an SV40 poly (A) termination signal, a U6 promoter, a sgRNA-3'-ITR inverted repeat sequence. The AAV-CRISPRi virus system can realize efficient and long-acting transcriptional inhibition on the ASIC3 gene in dorsal root ganglion neurons, and provides a novel treatment strategy with tissue targeting, lasting effect and no addiction risk for relieving inflammatory and neuropathic chronic pain.

The invention discloses a construction method and application of an AAV single-molecule sequencing library and a kit, and belongs to the technical field of biological medicine detection. In order to solve the technical problem that ligase steric hindrance is caused by a secondary structure of an AAV inverted repeat (ITR), a Cas9-RNP compound specifically targeting an ITR stem region (Stem Region) is used for carrying out directional cutting on a single-chain genome, a closed-loop hairpin structure is removed, and a blunt-end double-chain DNA handle of 10-50 bp is reserved at the tail end. In cooperation with a forced dissociation step of the enzyme, the method eliminates connection steric hindrance and significantly improves the linker connection efficiency. According to the method, double-end cyclization is not needed, sequencing can be achieved only through single-end connection, and therefore a complete genome, a truncated body and a covalent head-to-tail concatemer can be captured without bias, and the method can be used for accurately measuring the physical packaging capacity limit of the AAV carrier.

A method for improving the replication of a covalently closed circular plasmid is provided. The method includes providing a covalently closed circular plasmid having a Pol I-dependent origin of replication, and an insert including a structured DNA sequence selected from inverted repeat sequences, direct repeat sequences, homopolymeric repeat sequences, eukaryotic origins of replication or eukaryotic promoter enhancer sequences, wherein the structured DNA sequence is located at a distance of less than 1000 bp from the Pol I-dependent origin of replication in the direction of replication. The method also includes modifying the covalently closed circular recombinant molecule such that the Pol I-dependent origin of replication is replaced with a Pol III-dependent origin of replication, whereby the resultant Pol III-dependent origin of replication covalently closed circular plasmid has improved replication. An antibiotic marker free covalently closed circular recombinant DNA molecule is also provided.

An adeno-associated virus (AAV) complex platform is disclosed that includes an asymmetrically modified terminal inverted repeat (ITR). The AAV complex has an asymmetric ITR in which any one of two ITRs is modified, and thus has the advantages of increased productivity and transgene expression efficiency, and reduced genetic toxicity. Also disclosed are compositions comprising the adeno-associated virus complexes and gene therapy methods.

A method for producing recombinant adeno-associated virus particle preparations (rAAVp) is reported herein, comprising the steps of culturing mammalian cells containing expression cassettes for non-adeno-associated virus genes, adeno-associated virus rep genes, adeno-associated virus cap genes, adeno-associated virus E1A genes, adeno-associated virus E1B genes, adeno-associated virus E2A genes, adeno-associated virus E4orf6, and adeno-associated virus VA RNA genes located between two AAV terminal inverted repeat sequences (ITRs), and producing rAAVp therefrom, wherein the culturing is performed at a pH value between pH 7.4 and pH 7.6. The yield of rAAVp produced by culturing at a pH value between pH 7.4 and pH 7.6 is higher than that of rAAVp produced by culturing at a pH value between pH 7.0 and pH 7.2, and the percentage of perfect particles is higher for rAAVp produced by culturing at a pH value between pH 7.4 and pH 7.6 than for rAAVp produced by culturing at a pH value between pH 7.0 and pH 7.2.

The invention belongs to the technical field of biological medicine, and particularly relates to the field of gene therapy and pain therapy. The invention provides an sgRNA and AAV-CRISPRi virus system for targeted inhibition of TRPV1 gene, the AAV-CRISPRi virus system is a CRISPRi system based on AAV delivery, and the core functional structure sequence of the AAV-CRISPRi virus system is a 5 '-ITR inverted repeat sequence, an EF1 alpha core promoter, a ZIM3 transcription inhibition structural domain, a linker, a sadCas9-NLS nuclear localization signal-SV40 poly (A) termination signal-U6 promoter, and an sgRNA-3'-ITR inverted repeat sequence, according to the core functional structure, the miniaturization of a CRISPRi system is realized, and the infection success rate and the target gene inhibition efficiency are improved while the plasmid construction and AAV packaging costs are reduced. The AAV-CRISPRi virus system for targeted inhibition of the TRPV1 gene is used for preparing drugs for chronic pain, and has the characteristics of high efficiency, strong specificity and few side effects.

The present disclosure relates generally to ultra-minimum transposon inverted repeat (ITR) polynucleotides, compositions comprising the polynucleotides, and methods of delivering nucleic acids to cells ex vivo and in vivo, particularly therapeutic genes in vivo, using the compositions comprising the polynucleotides to treat genetic disorders or diseases.