4 results about "Direct repeat" patented technology

The present invention relates to a method for recording a transcriptome of a cell by: providing a test cell that includes a first transgene nucleic acid sequence encoding a fusion protein that is a reverse transcriptase polypeptide and a Cas1 polypeptide and a second transgene nucleic acid sequence encoding a Cas2 polypeptide, wherein the first transgene nucleic acid sequence and the second transgene nucleic acid sequence are under transcriptional control of an inducible promoter sequence, and a third transgene nucleic acid sequence including a CRISPR direct repeat (DR) sequence; wherein the CRISPR direct repeat sequence is specifically recognizable by a RT-Cas1-Cas2 complex formed by the expression products of the first transgene nucleic acid sequence and the second transgene nucleic acid sequence.

A CRISPR-Cas system and applications thereof are provided, also related to a Cas protein, a fusion protein and a guide polynucleotide. The Cas protein has at least 50% sequence identity compared to the sequence set forth in any one of SEQ ID NOs: 1-184, 426-428. The fusion protein comprises the Cas protein fused to a protein domain and / or a polypeptide tag. The guide polynucleotide comprises a direct repeat sequence having at least 70% sequence identity to any one of SEQ ID NOs: 185-199, 245-247, 287-303, 369 and 370-373, and a guide sequence engineered to hybridize to a target nucleic acid. The CRISPR-Cas system comprises a Cas protein having at least 90% sequence identity compared to the sequence set forth in any one of SEQ ID NOs: 1-184, 426-428, or a nucleic acid encoding thereof, and a guide polynucleotide or a nucleic acid encoding thereof.

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 RNA interference-based product and method for inhibiting the expression of the double homeobox 4 (DUX4) gene on human chromosome 4q35 are disclosed. The invention comprises the silencing of the Cas13 protein of RNA, wherein Cas13 is specifically targeted to a DUX4 region of interest using a sequence-specific guide RNA (gRNA). The recombinant adeno-associated virus of the invention is constructed with a Cas13 direct repeat sequence and delivers DNA encoding an inhibitory gRNA that knocks down DUX4 expression. The method is applicable to the treatment of muscular dystrophy, including but not limited to facial-scapulohumeral muscle dystrophy (FSHD), and other disorders, including cancer, associated with elevated DUX4 expression.