47 results about "Splice Site SNP" patented technology

The present invention provides synthetic 5′UTRs comprising a first polynucleotide fragment and a second polynucleotide fragment, wherein the first polynucleotide fragment comprises at least one splice site of a first eukaryotic gene, the second polynucleotide fragment comprises at least a portion of 5′ untranslated region of a second eukaryotic gene, and the first polynucleotide fragment is located 5′ of the second polynucleotide fragment. In one embodiment, the first polynucleotide fragment comprises the second intron of a sarcoplasmic / endoplasmic reticulum calcium ATPase gene and the second polynucleotide fragment comprises at least a portion of the 5′ untranslated region (5′UTR) of a eukaryotic casein gene. The synthetic 5′UTRs are useful for increasing the expression of a transgene when positioned between a promoter and a transgene within an expression vector. The present invention also provides vectors comprising synthetic 5′UTRs and methods for increasing the expression of a transgene using synthetic 5′UTRs.

Disclosed are methods and constructs for engineering circular RNA. Disclosed is a vector for making circular RNA, said vector comprising the following elements operably connected to each other and arranged in the following sequence: a.) a 5' homology arm, b.) a 3' group I intron fragment containing a 3' splice site dinucleotide, c.) optionally, a 5' spacer sequence, d.) a protein coding or non-coding region, e.) optionally, a 3' spacer sequence, f.) a 5' Group I intron fragment containing a 5' splice site dinucleotide, and g.) a 3' homology arm, said vector allowing production of a circular RNA that is translatable or biologically active inside eukaryotic cells. In another embodiment, the vector can comprise the 5' spacer sequence, but not the 3' spacer sequence. In yet another embodiment,the vector can comprise the 3' spacer sequence, but not the 5' spacer sequence. Also disclosed is a method for purifying the circular RNA produced by the vector and the use of nucleoside modifications in circular RNA produced by the vector.

The present invention relates to a method of modulating splice site selection, splicing and alternative, the method comprising the step of hybridizing an oligonucleotide-protein conjugate to a target pre-mRNA molecule in a cell or cell extract, wherein the oligonucleotide-protein conjugate comprises an oligonucleotide moiety which comprises at least two distinct sequence elements: (i) a nucleic acid sequence that is complementary to a specific region upstream of the splice site in the target pre-mRNA molecule; and (ii) an extension containing a protein binding site sequence element for covalently binding a protein; wherein the protein moiety comprises a protein capable of modulating splicing of the splice site upon binding with the protein binding site.

The invention discloses a probe for detecting expression abundance of circular RNA. The probe includes backsplicing sites at the 5' terminal of a donor exon and at the 3' terminal of an accepter exon of the circular RNA. The invention also discloses a gene chip containing the probe, and a method for detecting the expression abundance of the circular RNA by virtue of the gene chip, wherein the method mainly comprises the following steps: 1) extracting and purifying total RNA of a to-be-detected sample; 2) removing ribosome RNA; 3) enriching circular RNA; 4) conducting linear amplification to the enriched circular RNA and labeling fluorescence; 5) hybridizing a fluorescence-labeled product with a chip probe; and 6) washing and scanning the chip, and analyzing data. By designing the probe and the gene chip which are capable of achieving specific detection of the expression abundance of the circular RNA, the rapid and high-throughput detection of the expression abundance of the circular RNA can be achieved, so that shortcomings in an existing circular RNA detection technology are overcome.

Mutations that affect mRNA splicing often produce multiple mRNA isoforms containing different exon structures. Definition of an exon and its inclusion in mature mRNA relies on joint recognition of both acceptor and donor splice sites. The instant methodology predicts cryptic and exon skipping isoforms in mRNA produced by splicing mutations from the combined information contents and the distribution of the splice sites and other regulatory binding sites defining these exons. In its simplest form, the total information content of an exon, Ri,total, is the sum of the information contents of its corresponding acceptor and donor splice sites, adjusted for the self-information of the exon length. Differences between Ri,total values of mutant versus normal exons that are concordant with gene expression data demonstrate alterations in the structures and relative abundance of the mRNA transcripts resulting from these mutations.

Disclosed herein are point mutations in the LMNA gene that cause HGPS. These mutations activate a cryptic splice site within the LMNA gene, which leads to deletion of part of exon 11 and generation of a mutant Lamin A protein product that is 50 amino acids shorter than the normal protein. In addition to the novel Lamin A variant protein and nucleic acids encoding this variant, methods of using these molecules in detecting biological conditions associated with a LMNA mutation in a subject (e.g., HGPS, arteriosclerosis, and other age-related diseases), methods of treating such conditions, methods of selecting treatments, methods of screening for compounds that influence Lamin A activity, and methods of influencing the expression of LMNA or LMNA variants are also described. Oligonucleotides and other compounds for use in examples of the described methods are also provided, as are protein-specific binding agents, such as antibodies, that bind specifically to at least one epitope of a Lamin A variant protein preferentially compared to wildtype Lamin A, and methods of using such antibodies in diagnosis, treatment, and screening. Also provided are kits for carrying out the methods described herein.

The invention discloses an alternative splicing reporter vector. Two sections of intron sequences intron1 and intron2 are inserted into a reporter gene sequence of a reporter gene expression vector pEGFP-C1; each of the intron1 and the intron2 has a 5' splicing site, a branch site and a 3' splicing site. The invention also discloses a preparation method of the alternative splicing reporter vector.The intron1 and the intron2 are designed and synthesized according to a splicing mechanism of RNA (Ribonucleic Acid); the intron1 and the intron2 are inserted into a reporter sequence of the reportergene expression vector respectively and are used for detecting an alternative splicing event on a cell level and a living body level; the introns containing the splicing sites are inserted so that correct splicing of post-transcriptional mRNA (messenger Ribonucleic Acid) is facilitated.

The present invention provides synthetic 5′UTRs comprising a first polynucleotide fragment and a second polynucleotide fragment, wherein the first polynucleotide fragment comprises at least one splice site of a first eukaryotic gene, the second polynucleotide fragment comprises at least a portion of 5′ untranslated region of a second eukaryotic gene, and the first polynucleotide fragment is located 5′ of the second polynucleotide fragment. In one embodiment, the first polynucleotide fragment comprises the second intron of a sarcoplasmic / endoplasmic reticulum calcium ATPase gene and the second polynucleotide fragment comprises at least a portion of the 5′ untranslated region (5′UTR) of a eukaryotic casein gene. The synthetic 5′UTRs are useful for increasing the expression of a transgene when positioned between a promoter and a transgene within an expression vector. The present invention also provides vectors comprising synthetic 5′UTRs and methods for increasing the expression of a transgene using synthetic 5′UTRs.

The invention provides a method of producing multiple polypeptides, such as antibodies or antibody fragments, in a eukaryotic cell using a single expression vector. The expression vector is engineered to comprise two or more expression cassettes under the control of a single promoter wherein the expression cassettes have splice sites which allow for their alternative splicing and expression as two or more independent gene products at a desired ratio. Use of the vector for the efficient expression of recombinant antibodies in eukaryotic host cells is disclosed as well as the use of such antibodies in diagnostic and therapeutic applications.

The present invention provides a method for treating Usher's syndrome in a human subject including administering to the human subject an oligonucleotide having 8 to 30 linked nucleosides having a nucleobase sequence comprising a complementary region comprising at least 8 contiguous nucleobases complementary to a target region of equal length within exon 3 of an Usher RNA transcript.

The present invention relates to a method for the highly specific, targeted capture of regions of human genomes and transcriptomes from the blood, i.e. from cell free circulating DNA, exosomes, microRNA, circulating tumor cells, or total blood cells, to allow for the highly sensitive detection of mutation, expression, copy number, translocation, alternative splicing, and methylation changes using combined nuclease, ligation, polymerase, and massively parallel sequencing reactions. The method generates a collection of different circular chimeric single-stranded nucleic acid constructs, suitable for sequencing on multiple platforms. In some embodiments, each construct of the collection comprised a first single stranded segment of original genomic DNA from a host organism and a second single stranded synthetic nucleic acid segment that is linked to the first single stranded segment and comprises a nucleotide sequence that is exogenous to the host organism. These chimeric constructs are suitable for identifying and enumerating mutations, copy changes, translocations, and methylation changes. In other embodiments, input mRNA, IncRNA, or miRNA is used to generate circular DNA products that reflect the presence and copy number of specific mRNA's, IncRNA's splice-site variants, translocations, and miRNA.

Disclosed are compositions, reagents, methods, assays, and kits for quantitative and sensitive detection of target ribonucleic acid (RNA) sequences, particularly target RNA sequences that contain RNA spliced sites, hairpin stem-loops or other topological configurations of RNA secondary, tertiary, and quaternary structure, as well as linear RNA sequences. In some embodiments, fluorogenic modified-backbone oligonucleotide probes are employed for specific sequence hybridization across the target site, followed by chemical autoligation to produce a fluorescent molecule within the ligated probes. The autoligation detection reaction is performed isothermally or through thermocycling.

The present invention relates to a method of modulating splice site selection, splicing and alternative, the method comprising the step of hybridizing an oligonucleotide-protein conjugate to a target pre-mRNA molecule in a cell or cell extract, wherein the oligonucleotide-protein conjugate comprises an oligonucleotide moiety which comprises at least two distinct sequence elements: (i) a nucleic acid sequence that is complementary to a specific region upstream of the splice site in the target pre-mRNA molecule; and (ii) an extension containing a protein binding site sequence element for covalently binding a protein; wherein the protein moiety comprises a protein capable of modulating splicing of the splice site upon binding with the protein binding site.

Through targetting splicing sites of eukaryocyte genomes, the signal-noise ratio through negative heredity screening can be increased.

The current invention provides peptide nucleic acid derivatives targeting the 3' splice site of exon 4 in the human SCN9A pre-mRNA. The peptide nucleic acid derivatives potently induce SCN9A mRNA splice variant(s) lacking the SCN9A exon 4 in cells, and are useful to safely treat pains or conditions involving Nav1.7 activity.

There is provided SSOs targeting the region of HRAS, KRAS, and HRAS exon 2 that harbors the activating mutations and which harbors ESE activity. Moreover, there is provided SSOs targeting the 3′- and 5′-splice sites. The SSOs targeting the 5′ splice site sequence of HRAS exon 2, the 3′ splice site sequence of KRAS exon 2 and the 3′ splice site sequence of NRAS exon 2, as well as SSOs that targets ESEs in a conserved part of exon 2 in the HRAS, KRAS and NRAS exon 2 sequences can induce complete or nearly complete exon 2 skipping in cancer cell lines. This results in growth and proliferation inhibition and concomitantly in death of cancer cells. Therefore this invention is directed towards treatment of cancerous diseases and other conditions where RAS signaling is involved.

The present invention provides the peptide nucleic acid derivative which targets 5' splice site of the human ACC2 pre-mRNA "exon 12". The peptide nucleic acid derivatives in the present invention strongly induce splice variants of the human ACC2 mRNA in cell and are very useful to treat conditions or disorders of skin aging associated with the human ACC2 protein.

The invention discloses a method and device for detecting somatic cell SNP. The method comprises the steps of S1, comparing DNA sequencing data of somatic cells with reference genome data to obtain data of all SNP loci; S2, screening out SNP loci which occur on splicing loci and SNP loci which occur on exons and cause nonsynonymous mutation of amino acid to obtain a first SNP locus set; S3, removing SNP loci with the thousand-people mutation frequency higher than 5% in the first SNP locus set to obtain a second SNP locus set; S4, screening out SNP loci which are resources of somatic cell mutation from the second SNP locus set to obtain a third SNP locus set; S5, screening out SNP loci with the frequency of supporting sequences lower than 75% from the third SNP locus set to obtain SNP loci containing somatic cell SNP. By means of the method, somatic cell mutation can be detected through simple samples, and therefore the cost is lowered.