The present invention is directed to a nucleic acid construct comprising a polynucleotide operably linked to one or more control sequence that directs the expression of the polynucleotide in a host cell, wherein at least one control sequence comprises a promoter sequence of an operon comprising a secA gene or a functional fragment or functional variant thereof and wherein said promoter sequence is heterologous to the polynucleotide. In a further embodiment, the invention is directed to an expression vector and a host cell comprising the nucleic acid construct comprising the promoter sequence described herein and a method of expressing a polynucleotide in a host cell.

The present invention enables simultaneous and stable expression of a plurality of foreign genes by using a stealthy RNA gene expression system that is a complex that does not activate the innate immune mechanism and is formed from an RNA-dependent RNA polymerase, a single-strand RNA binding protein, and negative-sense single-strand RNAs including the following (1) to (8): (1) a target RNA sequence that codes for any protein or functional RNA; (2) an RNA sequence forming a noncoding region and derived from mRNA expressed in animal cells; (3) a transcription initiation signal sequence recognized by the RNA-dependent RNA polymerase; (4) a transcription termination signal sequence recognized by the polymerase; (5) an RNA sequence containing a replication origin recognized by the polymerase; (6) an RNA sequence that codes for the polymerase and of which codons are optimized for the species from which an introduction target cell is derived; (7) an RNA sequence that codes for a protein for regulating the activity of the polymerase and of which codons are optimized for the species from which the introduction target cell is derived; and (8) an RNA sequence that codes for the single-strand RNA binding protein and of which codons are optimized for the species from which the introduction target cell is derived.

Methods of assembling modified adenoviruses, libraries of adenoviral gene modules and compositions thereof are provided herein.

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 noncoding 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.

Methods of assembling modified adenoviruses, libraries of adenoviral gene modules and compositions thereof are provided herein.

The present disclosure discloses polynucleotide sequences that can be used to regulate gene expression in plants. Terminator sequences from Sorghum bicolor and Oryza sativa that are functional in plants are disclosed. Nucleic acid molecules, recombinant expression constructs, plants and seed comprising these terminator sequences are further disclosed.

Simultaneous expression of a plurality of foreign genes by using a stealthy RNA gene expression system that is a complex that does not activate the innate immune mechanism and is formed from an RNA-dependent RNA polymerase, a single-strand RNA binding protein, and negative-sense single-strand RNAs including the following (1) to (8): (1) a target RNA sequence that codes for any protein or functional RNA; (2) an RNA sequence forming a noncoding region and derived from mRNA; (3) a transcription initiation signal sequence recognized by the RNA-dependent RNA polymerase; (4) a transcription termination signal sequence recognized by the polymerase; (5) an RNA sequence containing a replication origin recognized by the polymerase; (6) an RNA sequence that codes for the polymerase; (7) an RNA sequence that codes for a protein for regulating the activity of the polymerase; and (8) an RNA sequence that codes for the single-strand RNA binding protein.

The present application describes structures, systems, and methods for modeling and analysis of single-strand break (SSB) signaling and repair in a cell-free system. Also provided are methods of making the SSB structures and SSB signaling and repair systems. Methods and systems for identifying modulators of DNA damage response (DDR) activity for SSB repair are also described as well as methods of inhibiting SSB repair.

The present invention enables simultaneous and stable expression of a plurality of foreign genes by using a stealthy RNA gene expression system that is a complex that does not activate the innate immune mechanism and is formed from an RNA-dependent RNA polymerase, a single-strand RNA binding protein, and negative-sense single-strand RNAs including the following (1) to (8): (1) a target RNA sequence that codes for any protein or functional RNA; (2) an RNA sequence forming a noncoding region and derived from mRNA expressed in animal cells; (3) a transcription initiation signal sequence recognized by the RNA-dependent RNA polymerase; (4) a transcription termination signal sequence recognized by the polymerase; (5) an RNA sequence containing a replication origin recognized by the polymerase; (6) an RNA sequence that codes for the polymerase and of which codons are optimized for the species from which an introduction target cell is derived; (7) an RNA sequence that codes for a protein for regulating the activity of the polymerase and of which codons are optimized for the species from which the introduction target cell is derived; and (8) an RNA sequence that codes for the single-strand RNA binding protein and of which codons are optimized for the species from which the introduction target cell is derived.

Adeno-associated virus (AAV) Clade F vectors or AAV vector variants (relative to AAV9) for precise editing of the genome of a cell and methods and kits thereof are provided. Targeted genome editing using the AAV Clade F vectors or AAV vector variants provided herein occurred at frequencies that were shown to be 1,000 to 100,000 fold more efficient than has previously been reported. Also provided are methods of treating a disease or disorder in a subject by editing the genome of a cell of the subject via transducing the cell with an AAV Clade F vector or AAV vector variant as described herein and further transplanting the transduced cell into the subject to treat the disease or disorder of the subject. Also provided herein are methods of treating a disease or disorder in a subject by in vivo genome editing by directly administering the AAV Clade F vector or AAV vector variant as described herein to the subject.