The disclosure relates to compositions comprising a bean yellow dwarf virus (BeYDV) vector that enables cell-to-cell movement of a transgene on a mastrevirus vector using a movement protein and nuclear shutting protein from begomovrius. Such compositions enable transformation of plants using reduced amounts of agrobacterium.

The invention is based on an expression enhancer sequence derived from the RNA-2 genome segment of a bipartite RNA virus, in which a target initiation site in the RNA-2 genome segment has been mutated. Deletion of appropriate start codons upstream of the main RNA2 translation initiation can greatly increase in foreign protein accumulation without the need for viral replication. Also provided are methods, vectors and systems, including the hyper-translatable Cowpea Mosaic Virus (CPMV-HT) based protein expression system.

Systems and methods for gene targeting in plants, including systems and methods that include the use of geminiviruses and customizable endonucleases.

A method for increasing the efficiency of homologous recombination-based gene editing in a plant according to an embodiment of the present invention includes optimizing temperature and photoperiod conditions during tissue culture of plant cells, expressing factors required for homology-directed DNA repair (HDR) and factors for increasing the HDR efficiency by using a multiple replicon, or regulating the HDR pathway or non-homologous end joining (NHEJ) pathway.

A nucleic acid construct is provided. The construct comprises a tobacco rattle virus (TRV) sequence and a nucleic acid sequence encoding a single guide RNA (sgRNA) that mediates sequence-specific cleavage in a target sequence of a genome of interest, wherein the TRV sequence is devoid of a functional 2b sequence. Also provided are plant cells comprising the construct and uses of the construct in gene editing.

The present disclosure relates to plant-based recombinant protein production systems and their methods of production and use. The plant-based recombinant protein production system is a vector comprising a 5 UTR and a 3 UTR, wherein the 3 UTR comprises at least one terminator selected from the group consisting of: EU, IEU, NbACT3, NbACT617, NbACT567, Pin2, BDB501, BDB282, NbHSP, NbHSPb, Rep, RbcS, SIR, SIR 5/3, SIR 3, AtHSP, 35S, RepA, NOS, TMV, TNVD, PEMV, and BYDV. In certain implementations, the vector comprises two terminators in the 3 UTR, where the two terminators are fused to form a double terminator. For example, the double terminator comprises two members selected from the group consisting of: EU, IEU, NbACT3, NbACT617, NbACT567, Pin2, BDB501, BDB282, NbHSP, NbHSPb, Rep, RbcS, SIR, SIR 5/3, SIR 3, AtHSP, 35S, RepA, NOS, TMV, TNVD, PEMV, and BYDV. In some aspects, the vector further comprises a chromatin scaffold/matrix attachment region (MAR) that is downstream of the terminators.

The invention is based on an expression enhancer sequence derived from the RNA-2 genome segment of a bipartite RNA virus, in which a target initiation site in the RNA-2 genome segment has been mutated. Deletion of appropriate start codons upstream of the main RNA2 translation initiation can greatly increase in foreign protein accumulation without the need for viral replication. Also provided are methods, vectors and systems, including the hyper-translatable Cowpea Mosaic Virus (CPMV-HT) based protein expression system.

Embodiments herein include engineered CRISPR-Cas effector proteins that comprise at least one modification compared to an unmodified CRISPR-Cas effector protein that enhances binding of the CRISPR complex to the binding site and/or alters editing preference as compared to wild type. In certain embodiments, the CRISPR-Cas effector protein is a Type V-B effector protein, e.g., C2c1. Embodiments also include viral vectors for delivery of CRISPR-Cas effector proteins, including C2c1. In certain embodiments, the vectors allow packaging of the CRISPR-Cas effector protein within a single vector. The disclosure also includes delivery vectors, constructs, and methods of delivering larger genes for systemic delivery.

Disclosed herein are viral vectors based on modifications of the Citrus Tristeza virus useful for transfecting citrus trees for beneficial purposes. Included in the disclosure are viral vectors including one or more gene cassettes that encode heterologous polypeptide s. The gene cassettes are positioned at desirable locations on the viral genome so as to enable expression while preserving functionality of the virus. Also disclosed are methods of transfecting plants and plants transfected with viral vector embodiments.

The invention is based on an expression enhancer sequence derived from the RNA-2 genome segment of a bipartite RNA virus, in which a target initiation site in the RNA-2 genome segment has been mutated. Deletion of appropriate start codons upstream of the main RNA2 translation initiation can greatly increase in foreign protein accumulation without the need for viral replication. Also provided are methods, vectors and systems, including the hyper-translatable Cowpea Mosaic Virus (CPMV-HT) based protein expression system.