The present invention relates to the field of genetic engineering and more particularly to nucleic acid editing and genome modification. The present invention provides an isolated Cas protein or polypeptide fragment thereof having an amino acid sequence of SEQ ID NO: 1 or a sequence of at least 77% identity therewith. The Cas protein or polypeptide is capable of binding, cleaving, marking or modifying a double stranded target polynucleotide at a temperature in the range 30° C. and 100° C. inclusive. The invention further provides isolated nucleic acid molecules encoding the Cas9 nucleases, expression vectors and host cells. The invention also provides PAM sequences recognized by the Cas protein or polypeptide, The Cas9 nucleases disclosed herein provide novel tools for genetic engineering at elevated temperatures and are of particular value in the genetic manipulation of thermophilic organisms; particularly microorganisms.

Provided are a human PD-1 knockdown siRNA, a recombinant expression CAR-T vector, a preparation method thereof, and an application of the same. A PD-1 knockdown siRNA expression cassette and an siRNA expression product thereof can be applied to a CAR-T therapy of multiple myeloma (MM) for eliminating or alleviating a tumor immune escape mechanism, and in the suppression of an immune escape mechanism in a CAR-T therapy of a tumor, such as pancreatic cancer, brain glioma, and myeloma.

Nucleic acid molecules such as shRNA clusters and artificial miRNA clusters are disclosed, Also disclosed are methods of use, compositions, cells, viral particles, and kits relating to the nucleic acid molecules disclosed herein. The disclosure provides, at least in part nucleic acid molecules such as shRNA clusters encoding shRNA-like molecules and artificial miRNA clusters encoding modified pri-miRNA-like molecules. The shRNA clusters and artificial miRNA clusters disclosed herein can be used, for example, to produce artificial RNA molecules, e.g., RNAi molecules. Cells, viral particles, compositions (e.g., pharmaceutical compositions), kits, and methods relating to the nucleic acid molecules, e.g., shRNA clusters and artificial miRNA clusters, are also disclosed. The nucleic acid molecules (e.g., shRNA clusters and artificial miRNA clusters), artificial RNA molecules (e.g., RNAi molecules), cells, viral particles, compositions (e.g., pharmaceutical compositions), and kits and methods disclosed herein can be used to treat or prevent a disease, e.g., HIV infection and/or AIDS.

This disclosure describes a composition and method of magenitic nanoparticles (MNP) that are bound to a baculovirus (BV). The MNP-BV can be systemically administered to a patient, and a strong magnetic field applied to the target btissue, thus allowing uptake and expression only in the target tissue. Off-target effects are not seen because the MNP-BC is inactivated by the complement system outside of the magnetic field.

Compositions and methods are provided for measuring population size for a plurality of clonal cell populations in the same individual, e.g., for measuring tumor size for a plurality of clonally independent tumors within the same individual. A subject method can include: (a) contacting an individual with a plurality of cell markers that are heritable and distinguishable from one another, to generate a plurality of distinguishable lineages of heritably marked cells; (b) after sufficient time has passed for the heritably marked cells to undergo at least one round of division, detecting and measuring quantities of at least two of the plurality of cell markers present in the contacted tissue, thereby generating a set of measured values: and (c) using the set of measured values to calculate the number of heritably marked cells that are present (e.g., for at least two of the distinguishable lineages of heritably marked cells).

The present invention includes a live attenuated virus and methods of making the same comprising an isolated virus comprising a viral genome that expresses one or more viral antigens; and one or more exogenous species-specific microRNAs inserted into the viral genome and expressed thereby, wherein the species-specific microRNAs are ubiquitously expressed in a viral target species cell but not in a viral propagation cell.

Methods and materials for improved delivery of dsRNA are presented. In particular, methods for controlling an insect pest by stably delivering large quantities of dsRNA to the insect pest are provided. Compositions comprising a bacterium containing a large quantity of dsRNA targeting an insect gene in which the dsRNA exhibits increased stability and activity are also provided.

The present disclosure relates to RNA interference-based methods for inhibiting the expression of the dystrophia myotonia protein kinase (DMPK) gene. Recombinant adeno-associated viruses of the disclosure deliver DNAs encoding inhibitory RNAs that knock down the expression of DMPK or interfere with the expression of the CTG repeat associated with myotonic dystrophy type-1 (DM1). The methods have application in the treatment of myotonic dystrophies, including DM1, and other disorders associated with aberrant DMPK expression.

The present invention relates to methods for shifting the splicing profile of the DUX4 gene, a double homeobox gene on human chromosome 4q35. Recombinant adeno-associated viruses of the invention deliver DNAs encoding U7-based small nucle-ar RNAs to induce DUX4 exon- skipping and the expression of shortened forms of DUX4. The methods have application in the treatment of muscular dystrophies such as facioscapulohumeral muscular dystrophy.

The present invention provides recombinant filamentous fungal host cells producing one or more secreted polypeptide of interest, said cells comprising in their genome at least one nucleic acid construct comprising a first polynucleotide encoding a signal peptide operably linked in translational fusion to a second polynucleotide encoding the polypeptide of interest, wherein the first polynucleotide is heterologous to the second polynucleotide, wherein the first polynucleotide is a polynucleotide having at least 70% sequence identity with SEQ ID NO:1 or a polynucleotide encoding a signal peptide having at least 70% sequence identity with SEQ ID NO:2, as well as methods of producing one or more secreted polypeptide of interest.