16 results about "Splice Site SNP" patented technology

A method and compound for regulating SCN1A gene mRNA and protein expression. Provided is a use of a guide agRNA in preparation of a drug for altering expression of a target protein in a cell of a subject. The cell has a precursor mRNA encoding the target protein, wherein the precursor mRNA comprises an intron, an exon flanking a 5' splice site of the intron, and an exon flanking a 3' splice site or a pseudo 3' splice site of the intron; the guide agRNA is capable of binding to the precursor mRNA to form a structure capable of recruiting ADAR in the cell, thereby forming a complex containing an ADAR protein; and the target protein is a Nav1.1 protein. By regulating alternative splicing of a target gene pre-mRNA, the functional RNA of the target gene and the function or expression of the target protein are affected, thereby achieving the objective of disease treatment.

A method for regulating the mRNA and protein expression of an SCN1A gene, and a compound. Provided is a use of an antisense oligonucleotide guide RNA in the preparation of a drug for altering the expression of a target protein in a cell of a subject. The cell has a pre-mRNA encoding the target protein, wherein the pre-mRNA comprises an intron, an exon flanking a 5' splice site of the intron, and an exon flanking a 3' splice site or pseudo 3' splice site of the intron; the antisense oligonucleotide guide RNA can bind to the pre-mRNA to form a structure capable of recruiting ADAR in a cell, thereby forming a complex containing an ADAR protein; and the target protein is an Nav1.1 protein. That is, the function or expression of the functional RNA and target protein of a target gene is affected by regulating the alternative splicing of the pre-mRNA of the target gene, thereby achieving the objective of treating diseases.

The invention discloses a tumor tissue splicing recognition and neoantigen peptide screening method based on an ONT sequencing platform, and belongs to the technical field of tumor immunology. Comprising the following steps: extracting RNA from a tumor tissue, separating mRNA, constructing a sequencing library, and sequencing through an ONT sequencing platform to obtain original data; performing basic group identification on the original data to obtain fastq format data; comparing the data to a reference genome reconstruction transcript, identifying a variable splicing event based on a comparison result, and performing difference analysis; screening out variable splicing events with significant differences, and extracting novol transcript coding sequences related to the events; extracting a 9mer peptide fragment crossing a splicing site in a splicing boundary region of the transcript; performing MHC binding affinity scoring on the peptide fragments, and screening out candidate new antigen peptide fragments; and carrying out mass spectrum verification on the candidate new antigen peptide fragment, and screening the actually expressed peptide fragment. The method can be widely applied to tumor immunomarker screening, new antigen vaccine design and individualized treatment strategy development.

A method of preparing a circular RNA includes transcribing a vector to form a precursor RNA, in which the vector includes the following elements operably connected to each other and arranged in the following sequence: a) a 5′ element, b) a 3′ Group I self-splicing intron fragment containing a 3′ splice site dinucleotide, c) none or an element containing an internal ribosome entry site (IRES) and a protein coding region or an element containing a noncoding region, d) a 5′ Group I self-splicing intron fragment containing a 5′ splice site dinucleotide, and e) a 3′ element, in which 5′ element and 3′ element form a stable structure with a Gibbs free energy (ΔG) from −190 kcal / mol to −9.0 kcal / mol, provided that the stable structure is not a duplex with at least 95% base pairing between 5′ element and 3′ element, in which 3′ Group I self-splicing intron fragment and 5′ Group I self-splicing intron fragment form a self-cleaving and self-ligating RNA molecule, thereby generating circular RNA.

The invention relates to the technical field of plant genetic engineering, in particular to a gene down-regulation method based on regulation of precursor mRNA (messenger Ribonucleic Acid) splicing efficiency, which comprises the following steps: firstly, identifying at least one intron 5'terminal splicing site containing a GT sequence in a target gene; aiming at the site, designing a guide RNA (Ribonucleic Acid) sequence capable of targeting the downstream fourth or fifth nucleotide, and verifying the cleavage activity of the guide RNA sequence; constructing a gene editing vector by using the guide RNA sequence which is verified to be effective and a Cas9 nuclease coding gene; introducing the vector into a plant cell, carrying out genetic transformation and regeneration to obtain a transformed plant, and screening out a mutant which is subjected to site-directed editing at a target site through DNA (Deoxyribose Nucleic Acid) sequencing; and finally, extracting RNA (Ribonucleic Acid) of the mutant, reversely transcribing into cDNA (Complementary Deoxyribonucleic Acid), and analyzing transcript change through PCR (Polymerase Chain Reaction) amplification and gel electrophoresis. According to the invention, stable, heritable and accurate knock-down of gene expression is realized, and a new tool is provided for essential gene function research and crop character improvement.

A member of the Solanum species, such as a tomato (Solanum lycopersicum) plant, is provided that exhibits a jointless phenotype, wherein the plant comprises a Jointless-2 (j2) allele that is defined by the removal or disruption of a splice site within the SlMBP21 (Solyc12g038510) gene The removal or disruption of a splice site results in the complete or partial loss of at least one exon in an mRNA transcribed from a SlMBP21 gene (Solyc12g038510). More particularly, the removal or disruption of a splice site results in the complete or partial loss of exon 5 in an mRNA transcribed from a SlMBP21 gene (Solyc12g038510). The removal or disruption of a splice site may be carried out using precision breeding techniques.

The present invention relates to a therapeutic oligonucleotide for use in treating a disease in a subject that is associated with or caused by a missense, nonsense or in-frame indel mutation in a gene of the genome of the subject, wherein the therapeutic oligonucleotide is capable of specifically hybridizing to the 5'-splice site, the 3'-splice site or an exonic splicing enhancer (ESE) of an out-of- frame exon in the gene with the missense, nonsense or in-frame indel mutation, thereby causing skipping of the out-of-frame exon.

The technology disclosed relates to splice site prediction and aberrant splicing detection. In particular, it relates to a splice site predictor that includes a convolutional neural network trained on training examples of donor splice sites, acceptor splice sites, and non-splicing sites. An input stage of the convolutional neural network feeds an input sequence of nucleotides for evaluation of target nucleotides in the input sequence. An output stage of the convolutional neural network translates analysis by the convolutional neural network into classification scores for likelihoods that each of the target nucleotides is a donor splice site, an acceptor splice site, and a non-splicing site.

The embodiment of the invention discloses a gene splicing site recognition method and system based on small molecule regulation, and the method comprises the steps: obtaining the gene expression quantity of each candidate gene sequence, so as to select a plurality of reliable gene sequences; obtaining a candidate pseudo exon insertion event of each reliable gene sequence under the treatment of each small molecule compound, and selecting a reliable pseudo exon insertion event from a plurality of candidate pseudo exon insertion events; training a pre-constructed deep learning model according to the reliable gene sequence and the reliable pseudo exon insertion event to obtain a splicing site recognition model; obtaining a target gene sequence, and obtaining a base position feature sequence corresponding to the target gene sequence; and inputting the base position feature sequence into a splicing site identification model to predict the probability that each base is a splicing site through the splicing site identification model so as to obtain a splicing site coordinate corresponding to the target gene sequence according to the probability, thereby improving the splicing site identification accuracy.

This disclosure provides a DNA construct comprising: (a) a start exon encoding a start codon; (b) a first frameshift exon downstream of the start exon, the first frameshift exon including a first receptor splice site, wherein inclusion or exclusion of the first frameshift exon in alternative splicing determines the downstream reading frame of a mature mRNA transcript; and (c) a first specific exon downstream of the first frameshift exon, the first specific exon including a second receptor splice site, a first reporter gene, and a second reporter gene, wherein the first reporter gene is located upstream or downstream of the second reporter gene, and wherein the first reporter gene and the second reporter gene are in different reading frames, wherein exclusion of the first frameshift exon causes the first reporter gene to be in the same reading frame as the start codon, and wherein inclusion of the first frameshift exon causes the second reporter gene to be in the same reading frame as the start codon.

A peptide nucleic acid derivative of Formula I is provided to tightly bind to a splice site within a pre-mRNA in a sequence specific manner. Given with excellent cell membrane permeability and strong affinity for RNA, the peptide nucleic acid derivative induces exon skipping in cells treated with the peptide nucleic acid at sub-femtomolar concentration as “naked” oligonucleotide. The compound shows therapeutic activity in subjects upon systemic administration even at 1 μg / Kg or less, and therefore is useful to treat a disease or symptom at affordable treatment cost.

The invention belongs to the field of bioinformatics and genomics, and discloses a method for rapidly and accurately predicting an intron splicing site of a target species. The method comprises the following steps: 1) obtaining a genome sequence and transcriptome sequencing data of a target species; 2) screening annotation genes and constructing a training set and a test set; 3) performing learning training on splicing site information by adopting a deep learning model architecture combining a convolutional neural network, a bidirectional long-short-term memory network and a self-attention structure; 4) performing bit-by-bit prediction on the input gene sequence by using the model obtained by training; and 5) judging the position of the splicing site according to the prediction score, and labeling and re-evaluating the abnormal or low-credibility splicing site. The calculation model provided by the invention can learn key sequence features at splicing sites, and can be extended to various species through transfer learning to realize high-precision introne boundary prediction. According to the method, the gene annotation efficiency can be remarkably improved, splicing abnormity caused by mutation can be accurately recognized, and the method has important application value for genomics research and gene function analysis and regulation.

The invention discloses snRNA for correcting protein C defects caused by exon-intron boundary mutation and application of the snRNA. Specific U1 small nuclear RNA (U1snRNA) and U7 small nuclear RNA (U7snRNA) are designed for protein C defects caused by abnormal mRNA splicing due to PROC gene exon-intron boundary mutation, so that recognition of 5'splicing sites is enhanced or recovered, and activated potential splicing sites are shielded. The small nuclear RNA is expressed through respective expression vectors, and corresponding U1snRNA or U7snRNA is correspondingly designed for each mutation. Cell experiment results show that the U1snRNA and the U7snRNA can remarkably recover correct splicing of abnormal PROC transcripts and improve the expression level and the functional activity of protein C. The invention provides a small nuclear RNA strategy based on RNA, which has potential gene therapy application value.

Provided herein are polynucleotides, vectors, complexes, compositions, systems, kits, methods and uses for enriching for gene editing in a cell. Some aspects of the disclosure relate to the use of a polynucleotide cassette comprising a coding sequence of a selection marker gene that is disrupted by a synthetic intron sequence to prevent gene expression of the selection marker. The synthetic intron further comprises a defective splice site that is correctable by a base edit such that when the splice site is corrected by a gene editor, the intron is removed by the endogenous slicing system of the cell and expression of the selection marker gene can occur. Methods described herein screen for the presence of active gene editors within the cell by subjecting the cell to the selection pressure of the selection marker.

The present disclosure features bifunctional oligonucleotides and related compositions that, inter alia, modulate nucleic CA acid splicing, e.g., splicing of a pre-mRNA, as well as methods of use thereof.

The present application relates to an RNA molecule, which contains: a 3' self-splicing intron fragment containing a 3' splice site, a target sequence, and a 5' self-splicing intron fragment containing a 5' splice site, with the elements being operably linked in sequence, wherein the 3' self-splicing intron fragment and the 5' self-splicing intron fragment can enable circularization of the RNA molecule and can be removed from the RNA molecule during the circularization process; the 3'-terminal sequence of the target sequence ends with a nucleotide having a U base; the 5' self-splicing intron fragment contains an internal guide sequence, and the 5'-terminal sequence of the internal guide sequence is reverse complementary to the 5'-terminal sequence of the target sequence; and the 3'-terminal sequence of the internal guide sequence starts with a nucleotide having a G base and is reverse complementary to the 3'-terminal sequence of the target sequence starting from the end thereof.