Abstract: The present invention provides a method of compiling a plant positive sense functional gene profile, a method of changing the phenotype or biochemistry of a plant, a method of determining a change in phenotype or biochemistry of a plant, and a method of determining the presence of a trait in plant. The methods comprise expressing transiently a nucleic acid sequence of a plant into the cytoplasm of a host plant in a plus sense orientation to affect phenotypic or biochemical changes in the host plant. The nucleic acid sequence does not need to be isolated, identified or characterized prior to transfection into the host plant. A viral vector functional genomic screen has been developed to identify nucleotide sequences in transfected plants by systemically overproducing a new protein, or enhancing or suppressing the endogenous gene expression in a plus sense mechanism.

Abstract: The invention relates to &agr;-galactosidase truncated at the carboxy terminus and the production of enzymatically active recombinant human and animal lysosomal enzymes involving construction and expression of recombinant expression constructs comprising coding sequences of human or animal lysosomal enzymes in a plant expression system. The plant expression system provides for post-translational modification and processing to produce a recombinant gene product exhibiting enzymatic activity. The invention is demonstrated by working examples in which transgenic tobacco plants express recombinant expression constructs comprising human glucocerebrosidase nucleotide sequences. The invention is also demonstrated by working examples in which transfected tobacco plants express recombinant viral expression constructs comprising human &agr; galactosidase nucleotide sequences.

Abstract: The present invention provides a method of compiling a functional gene profile of an organism, a method of changing the phenotype or biochemistry of a plant, a method of determining a change in phenotype or biochemistry of a plant, a method of determining the presence of a trait in plant, and a method of isolating human cDNA. The methods comprise expressing transiently a nucleic acid sequence of a non-plant donor organism into a host plant by a viral vector to affect phenotypic or biochemical changes in the host plant. The present invention provides a method for discovering the presence of a new gene and determining its function and sequence in a donor organism such as human by transfecting a nucleic acid sequence of the donor organism into a host plant to knock out the endogenous gene expression.

Abstract: The present invention provides a method of compiling a functional gene profile of an organism, a method of changing the phenotype or biochemistry of a plant, a method of determining a change in phenotype or biochemistry of a plant, a method of determining the presence of a trait in plant, and a method of humanizing plant cDNA. The methods comprise expressing transiently a nucleic acid sequence of a non-plant donor organism into a host plant by a viral vector to affect phenotypic or biochemical changes in the host plant. The present invention provides a method for discovering the presence of a new gene and determining its function and sequence in a donor organism such as human by transfecting a nucleic acid sequence of the donor organism into a host plant to knock out the endogenous gene expression.

Abstract: The present invention provides a method of compiling a positive sense functional gene profile of an organism, a method of changing the phenotype or biochemistry of an organism, a method of determining the presence of a trait in an organism, and a method of humanizing a plant cDNA. The methods comprise expressing transiently a nucleic acid sequence of a donor organism into a host plant to affect phenotypic or biochemical changes in the host organism. Once the presence of a trait in a plant is identified by phenotypic or biochemical changes in the host plant, the nucleic acid insert in the cDNA clone or in the vector that results in the changes is then sequenced. The present invention provides a method for discovering new gene and its function in a donor organism such as human by transfecting a nucleic acid sequence of the donor organism into a host organism in a positive sense.

Abstract: The present invention provides a method of compiling a positive sense functional gene profile of an organism, a method of changing the phenotype or biochemistry of an organism, a method of determining a change in phenotype or biochemistry of an organism, and a method of determining the presence of a trait in an organism, and a method of isolating human cDNA. The methods comprise expressing transiently a nucleic acid sequence of a donor organism into a host plant to affect phenotypic or biochemical changes in the host organism. A viral vector functional genomic screen has been developed to identify nucleotide sequences in transfected plants by enhancing or supressing an endogenous gene expression in a positive sense mechanism, or by overexpressing a new protein. Once the presence of a trait in a plant is identified by phenotypic or biochemical changes in the host plant, the nucleic acid insert in the cDNA clone or in the vector that results in the changes is then sequenced.

Abstract: The present invention provides a method of compiling a plant functional gene profile, a method of changing the phenotype or biochemistry of a plant, a method of determining a change in phenotype or biochemistry of a plant, and a method of determining the presence of a trait in plant. The methods comprise expressing transiently a nucleic acid sequence of a plant into a host plant to affect phenotypic or biochemical changes in the host plant. A viral vector functional genomic screen has been developed to identify nucleotide sequences in transfected plants by systemically knocking out endogenous gene expression in an antisense mechanism. Once the presence of a trait in a plant is identified by phenotypic or biochemical changes in the host plant, the nucleic acid insert in the cDNA clone or in the vector that results in the changes is then sequenced. The present invention exemplifies that genes encoding GTP binding proteins in one plant can silence endogenous gene expression in a different plant.

Abstract: The present invention provides methods for rapidly determining the function of nucleic acid sequences by transfecting the same into a host organism to effect expression. Phenotypic and biochemical changes produced thereby are then analyzed to ascertain the function of the nucleic acids which have been transfected into the host organism. The invention also provides methods for silencing endogenous genes by transfecting hosts with nucleic acid sequences to effect expression of the same. The present invention also provides methods for selecting desired functions of RNAs and proteins by the use of virus vectors to express libraries of nucleic acid sequence variants. Moreover, the present invention provides methods for inhibiting an endogenous protease of a plant host.

Abstract: The present invention provides a method of compiling a plant functional gene profile, a method of changing the phenotype or biochemistry of a plant, a method of determining a change in phenotype or biochemistry of a plant, and a method of determining the presence of a trait in plant. The methods comprise expressing transiently a nucleic acid sequence of a plant into a host plant to affect phenotypic or biochemical changes in the host plant. A viral vector functional genomic screen has been developed to identify nucleotide sequences in transfected plants by systemically knocking out endogenous gene expression in an antisense mechanism. Once the presence of a trait in a plant is identified by phenotypic or biochemical changes in the host plant, the nucleic acid insert in the cDNA clone or in the vector that results in the changes is then sequenced. The present invention exemplifies that genes encoding GTP binding proteins in one plant can silence endogenous gene expression in a different plant.

Abstract: The present invention provides a method of compiling a plant functional gene profile, a method of changing the phenotype or biochemistry of a plant, a method of determining a change in phenotype or biochemistry of a plant, a method of determining the presence of a trait in plant, and a method of increasing grain crop. The methods comprise expressing transiently a nucleic acid sequence of a plant into a host plant to affect phenotypic or biochemical changes in the host plant. A viral vector functional genomic screen has been developed to identify nucleotide sequences in transfected plants by systemically knocking out endogenous gene expression in an antisense mechanism. Once the presence of a trait in a plant is identified by phenotypic or biochemical changes in the host plant, the nucleic acid insert in the cDNA clone or in the vector that results in the changes is then sequenced.

Abstract: The invention provides novel genetic constructions for the expression of inhibitory RNA in the cytoplasm of eukaryotic cells. The inhibitory RNA may be an anti-sense RNA or a co-suppressor RNA, and functions to reduce the expression of a gene of interest in the target cell. The genetic constructions of the invention are capable of replicating in the cytoplasm of a eukaryotic cell and comprise a promoter region in functional combination with an encoding polynucleotide. The genetic constructions may be designed so as to replicate in the cytoplasm of plant cells, yeast cells, and mammalian cells. When the eukaryotic cell of interest is a plant cell, the genetic construction is preferably derived from a plant RNA virus. Plant RNA virus derived genetic constructions may employ a plant virus subgenomic promoter, including subgenomic promoters from tobamoviruses in functional combination with the RNA encoding region.

Abstract: This invention is directed to a monopartite RNA viral vector comprising modified tobravirus RNA-1 comprising an inserted foreign RNA sequence. This invention is also directed to a bipartite RNA viral vector derived from a tobravirus, wherein the vector comprises one or more foreign RNA sequences. The invention is directed to a method of silencing one or more endogenous plant host genes and a method of simultaneously silencing a plant host gene and expressing a foreign gene in a plant host. Such methods comprise infecting a plant host with a bipartite vector comprising modified tobravirus RNA-1 and RNA-2. The invention is further directed to a method of compiling a plant functional gene profile, a method of changing the phenotype or biochemistry of a plant host, and a method of determining the presence of a trait in a plant host, using a monopartite or bipartite viral vector derived from a tobravirus.

Abstract: The present invention provides a method for enhancing the production of RNAs or proteins in a plant host using either non-native 5′ untranslated sequences or artificial leader sequences. Preferably, commercially useful proteins, polypeptides, or fusion products thereof are produced, such as, enzymes, antibodies, hormones, pharmaceuticals, vaccines, pigments, anti-microbial polypeptides, and the like. The non-native 5′ untranslated enhancers may also be effective in many different types of transcription or translation systems, such as bacterial and animal systems.

Abstract: The present invention provides a method of compiling a plant functional gene profile, a method of changing the phenotype or biochemistry of a plant, a method of determining a change in phenotype or biochemistry of a plant, and a method of determining the presence of a trait in plant. The methods comprise expressing transiently a nucleic acid sequence of a plant into a host plant to affect phenotypic or biochemical changes in the host plant. A viral vector functional genomic screen has been developed to identify nucleotide sequences in transfected plants by systemically knocking out endogenous gene expression in an antisense mechanism. Once the presence of a trait in a plant is identified by phenotypic or biochemical changes in the host plant, the nucleic acid insert in the cDNA clone or in the vector that results in the changes is then sequenced. The present invention exemplifies that genes encoding GTP binding proteins in one plant can silence endogenous gene expression in a different plant.

Abstract: The invention relates to -galactosidase truncated at the carboxy terminus and the production of enzymatically active recombinant human and animal lysosomal enzymes involving construction and expression of recombinant expression constructs comprising coding sequences of human or animal lysosomal enzymes in a plant expression system. The plant expression system provides for post-translational modification and processing to produce a recombinant gene product exhibiting enzymatic activity. The invention is demonstrated by working examples in which transgenic tobacco plants express recombinant expression constructs comprising human glucocerebrosidase nucleotide sequences. The invention is also demonstrated by working examples in which transfected tobacco plants express recombinant viral expression constructs comprising human &agr; galactosidase nucleotide sequences.

Abstract: The present invention relates to a method for conferring herbicide, pest, and disease resistance in plant hosts. Specifically, the present invention employs transient viral expression vectors to express proteins or enzymes conferring resistance in plant hosts. In addition, a library of nucleotide sequence variants in a sense or antisene orientation may be used to determine the targets of an herbicide or pathogen and to screen suitable viral nucleic acids for herbicide, pest, and disease resistance.

Abstract: The invention provides novel genetic constructions for the expression of inhibitory RNA in the cytoplasm of eukaryotic cells. The inhibitory RNA may be an anti-sense RNA or a co-suppressor RNA, and functions to reduce the expression of a gene of interest in the target cell. The genetic constructions of the invention are capable of replicating in the cytoplasm of a eukaryotic cell and comprise a promoter region in functional combination with an encoding polynucleotide. The genetic constructions may be designed so as to replicate in the cytoplasm of plant cells, yeast cells, and mammalian cells. When the eukaryotic cell of interest is a plant cell, the genetic construction is preferably derived from a plant RNA virus. Plant RNA virus derived genetic constructions may employ a plant virus subgenomic promoter, including subgenomic promoters from tobamoviruses in functional combination with the RNA encoding region.

Abstract: The present invention relates to a method for conferring herbicide, pest, and disease resistance in plant hosts. Specifically, the present invention employs transient viral expression vectors to express proteins or enzymes conferring resistance in plant hosts. In addition, a library of nucleotide sequence variants in a sense or antisene orientation may be used to determine the targets of an herbicide or pathogen and to screen suitable viral nucleic acids for herbicide, pest, and disease resistance.

Abstract: The invention provides novel genetic constructions for the expression of inhibitory RNA in the cytoplasm of eukaryotic cells. The inhibitory RNA may be an anti-sense RNA or a co-suppressor RNA, and functions to reduce the expression of a gene of interest in the target cell. The genetic constructions of the invention are capable of replicating in the cytoplasm of a eukaryotic cell and comprise a promoter region in functional combination with an encoding polynucleotide. The genetic constructions may be designed so as to replicate in the cytoplasm of plant cells, yeast cells, and mammalian cells. When the eukaryotic cell of interest is a plant cell, the genetic construction is preferably derived from a plant RNA virus. Plant RNA virus derived genetic constructions may employ a plant virus subgenomic promoter, including subgenomic promoters from tobamoviruses in functional combination with the RNA encoding region.

Abstract: Two novel genomic clones, ZZA1 and ZZA2, were isolated which encode for Pichia pastoris alcohol oxidase isozymes. The 5' non-coding region of ZZA1 contains common structural features involved in the transcription and translation of eukaryotic genes. Comparison of the nucleotide sequences of the ZZA1 and AOX1 51 noncoding regions showed that they are 66% similar to each other.The rice .alpha.-amylase gene OS103 was placed under the transcriptional control of the ZZA1 promoter. The nucleotide sequences of ZZA1 and other methanol-regulated promoters were analyzed. A highly conserved sequence (TTGNNNGCTTCCAANNNNNTGGT) (SEQ ID NO:2) was found in the 51 flanking region. A yeast strain containing the ZZA1 -OS103 fusion and secreting biologically active a-amylase into the culture media while converting starch to ethanol was produced. The ZZA1 and ZZA2 regulatory sequences may be used to contol the expression of other heterologous proteins in multiple yeast species.