The invention relates to the field of genetic engineering tools for gene expression in plants. Specifically, the invention concerns modified Foxtail Mosaic Virus (FoMV) vectors comprising polynucleotide sequences which are capable of driving expression of a gene of interest in a plant host. Accordingly, the invention concerns FoMV-based expression vectors comprising said polynucleotides, compositions comprising modified FoMV vectors, methods of generating gene expression in plants infected with the modified FoMV vectors. The expression vectors, compositions, plants and methods of the present invention find application in many fields of biotechnology, including, for example, gene characterization, protein production and agricultural biotechnology.

The invention provides a method of producing a potexviral vector for expressing a protein of interest in a plant, comprising producing a second heterologous nucleic acid comprising a second ORF encoding said protein and having, in the second ORF, an increased GC-content compared to a first ORF encoding said protein in a first heterologous nucleic acid, and providing said potexviral vector comprising the following segments: (i) a nucleic acid sequence segment encoding a potexviral RNA-dependent RNA polymerase, (ii) a nucleic acid sequence comprising or encoding a potexviral triple-gene block, and (iii) said second heterologous nucleic acid or a portion thereof comprising said second ORF.

Disclosed herein are methods and exemplary compositions associated with virus purification, antigen purification, and conjugation of virus and proteins (e.g., antigen) to form vaccines for delivery of immunological and other therapeutic agents, exemplary aspects of which may include harvesting viral and antigenic substances from source organisms; a purification platform comprising chemical separation and size-difference separation for the removal of contaminants, debris and impurities from the viral and protein (e.g. antigenic, including influenza hemagglutinin antigens) substances, as well as their concentration and collection; and a conjugation platform providing activation of the virus at a pH that increases binding rate and binding propensity between the virus and the protein, wherein embodiments related to the conjugation platform include controlling the ratio of virus to protein.

Disclosed herein are methods and exemplary compositions associated with virus purification, antigen purification, and conjugation of virus and proteins (e.g., antigen) to form vaccines for delivery of immunological and other therapeutic agents, exemplary aspects of which may include harvesting viral and antigenic substances from source organisms; a purification platform comprising chemical separation and size-difference separation for the removal of contaminants, debris and impurities from the viral and protein (e.g. antigenic, including influenza hemagglutinin antigens) substances, as well as their concentration and collection; and a conjugation platform providing activation of the virus at a pH that increases binding rate and binding propensity between the virus and the protein, wherein embodiments related to the conjugation platform include controlling the ratio of virus to protein.

Disclosed herein are viral vectors suitable for transfection into woody trees for purposes of delivering and expressing beneficial genes. Specifically exemplified herein are vectors for transfecting citrus trees. The vectors allow for the expression of useful proteins, such as those that can protect the tree from disease. Specifically exemplified herein are methods of transfecting woody trees that allow multiple applications of vectors while avoiding superinfection exclusion.

An object of the present invention is induction of CD4-positive T cells from pluripotent stem cells. This object is achieved by production of CD4-positive T cells by introducing a CD4 gene or a gene product thereof into T cells induced from pluripotent stem cells.

The invention concerns the targeted genomic modification of a plant cell, preferably a meristem cell. More in particular, the invention pertains to a vector expressing a coding RNA, wherein the coding RNA comprises a sequence encoding a CRISPR-nuclease and a mobile element, wherein the mobile element enables intercellular translocation of the coding RNA, preferably intercellular translocation to a meristem cell. The invention further concerns an editing RNA comprising the coding RNA and further comprising a guide RNA.

The present disclosure provides compositions and methods for editing a target site of a plant genome by delivery of functional editing components using modified tobacco mosaic virus (mTMV). The methods disclosed herein can be used to deliver a gene editing system, such as a DNA endonuclease, to a tobacco plant cell for modification of a target site of the plant genome. Further, the methods and compositions disclosed herein provide for production of a RNA molecule encoding a meganuclease in vitro prior to delivery of the RNA to a plant cell. After introduction of the nucleic acid molecule encoding a functional editing component and subsequent expression of the functional editing components, the plant can be cultured and allowed to produce seeds having an edit at a genomic target site. The seeds can then undergo embryo rescue and be cultured to produce a modified plant without heterologous genetic material.

The present invention relates to methods and compositions for enhancing expression from RNA expression vectores. The invention is based upon the observation that reducing the frequency of the dinucleotide CpG and UpA has a significant effect on expression from such vectores. Aspects of the invention include, amongst others, synthetic RNA vectores, virions, cells, methods of producing vaccines and methods of treatment or immunisation.

The present invention relates to kits comprising plus-sense single stranded RNA viral vectors, as well as mixtures of these vectors and uses thereof, and methods for producing in a plant, or plant tissue, or plant cell simultaneously two or more polypeptides using the kits and vectors.