19 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.

The application belongs to the technical field of biology, and discloses a kind of Myzus persicae densovirus, amplification and connector method and its application, Myzus persicae densovirus name is Myzus persicae densovirus 2, abbreviated as MpDV2, preserved in China typical culture preservation center, and the preservation number is V202281.The Myzus persicae densovirus MpDV2 provided in the application is a virus strain that has not been isolated and identified before and has pathogenicity to Myzus persicae, and the full-length sequence of the genome of the Myzus persicae densovirus MpDV2 (including the complete terminal inverted repeat sequence) is disclosed for the first time, and the influence of the virus infection on the growth and development of Myzus persicae and the population dynamics / growth.The Myzus persicae densovirus MpDV2 of the application significantly reduces the intrinsic growth rate (r), net reproduction rate (R0) and week-long growth rate (lambda) of Myzus persicae population, and prolongs the average generation time (T) of the population, and the MpDV2 infection has a significant influence on the growth and development of Myzus persicae and population growth.

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.

The present disclosure relates to circular, non-viral DNA vectors, compositions including one or more of the disclosed vectors, and methods for delivering and / or expressing one or more therapeutic genes (e.g., proteins) in mammals, e.g., human patients. In some embodiments, the present disclosure is directed to circular, non-viral DNA vectors, such as circular non-viral DNA vectors including at least two inverted repeat sequences, where the at least two inverted repeat sequences are separated by a non-repeated nucleotide sequence which is not part of the at least two inverted repeat sequences. In some embodiments, the disclosed circular, non-viral DNA vectors do not include a “DD element.” In some embodiments, the disclosed circular, non-viral DNA vectors do not include a “DD element,” but include at least a portion of a bacterial origin of replication.

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.

This invention discloses an RNAi recombinant vector resistant to both whitefly and tomato yellow leaf curl virus (TYLCV), its construction method, and its applications. The method involves constructing a DNAβ promoter associated with a geminivirus expressed in the phloem into the RNAi vector pCambia1391, obtaining a 1391::DNAβpro vector backbone. RT-PCR is used to amplify a partial fragment of the whitefly key gene BtACTB, a key region of TYLCV (including the intergenic spacer region for viral replication initiation, some coat proteins, and replication-related proteins), and its inverted repeat sequences. This constructs a recombinant vector with an inverted hairpin structure driven by the phloem promoter, simultaneously targeting key sequences of both whitefly and TYLCV. The constructed recombinant vector is then introduced into tomatoes via Agrobacterium-mediated transformation. Screening yields tomato plants that reduce whitefly survival and inhibit TYLCV infection.

This invention belongs to the field of biotechnology and discloses a high-yield rAAV plasmid system and its applications. The high-yield rAAV plasmid system of this invention includes a transgenic plasmid containing two terminal inverted repeat sequences, a packaging plasmid, a helper plasmid, and an shRNA expression cassette; the shRNA expression cassette includes a sequence capable of targeting the target gene in the transgenic plasmid. In the rAAV production process, the plasmid system of this invention inhibits the transcriptional expression of the target gene by expressing shRNA to target the coding region or target sequence of the target gene on the transgenic plasmid. The carefully designed target sequence, unlike coding regions of human and mammalian genes, can be broadly used in various transgenic plasmids, thereby increasing rAAV yield and significantly improving the rAAV production efficiency of mammalian cells, achieving the technical effects of improved rAAV druggability and reduced production costs.

The invention discloses insecticidal dsRNA synthesized according to KCC as well as a synthesis method and application of the dsRNA. The gene sequence of the dsRNA is shown as SEQ ID NO: 1 or at least comprises a gene sequence with 90% of identity of the sequence shown as SEQ ID NO: 1. After the compound is introduced into the body of a test insect through a micro-injection method, the accumulated death rate can reach 98.33%, and the lethal effect is remarkable. According to the invention, a recombinant expression vector containing an inverted repeat sequence of the gene is constructed, and high-efficiency expression and accumulation of dsRNA are realized through IPTG induction in escherichia coli. After the dsRNA-containing engineering bacteria are mixed with flour, test insects are fed, and the accumulative fatality rate can reach 58.33%. A metal organic framework (MOF) material is used for loading the obtained dsRNA, the stability and delivery efficiency of the dsRNA can be further improved, the final death rate can reach 65.56% after a test insect compound is fed, and an effective means is provided for application of nucleic acid biopesticide in prevention and control of stored grain pests.

To provide an adeno-associated virus (AAV) vector genome for altering, e.g., enhancing, expression of frataxin (FXN) in vitro and / or in vivo, including an engineered promoter.SOLUTION: Provided is an adeno-associated virus (AAV) vector genome comprising a 5' inverted terminal repeat (ITR), an engineered promoter, a payload region, and a 3' ITR, wherein the payload region encodes a frataxin (FXN) protein.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 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.

The present disclosure relates to circular, non-viral DNA vectors, compositions including one or more of the disclosed vectors, and methods for delivering and / or expressing one or more therapeutic genes (e.g., proteins) in mammals, e.g., human patients. In some embodiments, the present disclosure is directed to circular, non-viral DNA vectors, such as circular non-viral DNA vectors including at least two inverted repeat sequences, where the at least two inverted repeat sequences are separated by a non-repeated nucleotide sequence which is not part of the at least two inverted repeat sequences. In some embodiments, the disclosed circular, non-viral DNA vectors do not include a “DD element.” In some embodiments, the disclosed circular, non-viral DNA vectors do not include a “DD element,” but include at least a portion of a bacterial origin of replication.

Provided herein are double strand DNA molecules comprising inverted repeats, expression cassette and one or more restriction sites for nicking endonucleases, the methods of use thereof, and the methods of making therefor.

The invention provides novel DNA (deoxyribonucleic acid) binding proteins Aca11 and Aca13. The DNA binding proteins can be specifically bound with DNA by recognizing inverted repetitive sequences in the DNA. The invention also provides application of the DNA binding protein in transcriptional regulation of genes containing inverted repetitive sequences in cells. The invention also provides an application of the protein AcrIIA35 in inhibition of Cas9 gene editing or regulation in cells. The invention also provides a protein-protein interaction detection system based on Aca drive, and the system comprises a target protein with an Aca13 label, and a fluorescent DNA probe containing a conservative inverted repeat sequence capable of being specifically recognized by Aca13. As a powerful protein interaction research platform, the detection system provided by the invention provides a multifunctional tool for functional proteomics in CRISPR-Cas research.