1. Field of the Invention
Disclosed herein are recombinant DNA useful for providing enhanced traits to transgenic plants, seeds, pollen, plant cells and plant nuclei of such transgenic plants, methods of making and using such recombinant DNA, plants, seeds, pollen, plant cells and plant nuclei. Also disclosed are methods of producing hybrid corn seed comprising such recombinant DNA. Also disclosed are plants and seeds having an increased oil content or modified oil composition. All genetic resources disclosed herein were directly obtained from sources that are currently common to the United States; the ancestral sources of each specific genetic material is unknown.
2. Summary of the Invention
This invention provides recombinant DNA encoding or suppressing proteins with at least 95% identity to at least 95% of a sequence selected from SEQ ID NOs: 4-6. The invention further provides isolated polypeptides with at least 95% identity to at least 95% of a sequence selected from SEQ ID NOs: 4-6.
Another aspect of the invention further employs recombinant DNA for expression or suppression of proteins thereby imparting enhanced agronomic traits to the transgenic plants. Recombinant DNA in this invention is provided in a construct comprising a promoter that is functional in plant cells and that is operably linked to DNA that encodes or suppresses a protein having at least 95% identity to at least 95% of a sequence selected from the group consisting of SEQ ID NOs 4-6.
Other aspects of the invention are specifically directed to transgenic plant cells comprising the recombinant DNA of the invention, transgenic plants comprising a plurality of such plant cells, progeny transgenic seed, embryo and transgenic pollen from such plants. Such transgenic plants are selected from a population of transgenic plants regenerated from plant cells transformed with recombinant DNA and expressing or suppressing the protein(s) by screening transgenic plants in the population for an enhanced trait as compared to control plants that do not have said recombinant DNA, where the enhanced trait is selected from group of enhanced traits consisting of enhanced water use efficiency, enhanced cold tolerance, increased yield, enhanced nitrogen use efficiency, enhanced seed protein, enhanced seed oil and modified oil composition. In further embodiments, such transgenic plant cells include polynucleotide stacks which express or suppress multiple proteins of the invention. In a particularly specific embodiment of the invention, such transgenic plants comprise polynucleotide stacks encoding proteins which are at least 95% identical to at least 95% of SEQ ID NOs: 4-6.
In yet another aspect of the invention the plant cells, plants, seeds, embryo and pollen further comprise DNA expressing a protein that provides tolerance from exposure to an herbicide applied at levels that are lethal to a wild type plant cell. Such tolerance is especially useful not only as an advantageous trait in such plants but is also useful in a selection step in the methods of the invention. In aspects of the invention the agent of such herbicide is a glyphosate, dicamba, or glufosinate compound.
Yet other aspects of the invention provide transgenic plants which are homozygous for the recombinant DNA and transgenic seed of the invention from corn, soybean, cotton, canola, alfalfa, wheat or rice plants. In certain embodiments, for instance for practice of various aspects of the invention in Argentina, the recombinant DNA is provided in plant cells derived from corn lines that are and maintain resistance to the Mal de Rio Cuarto virus or the Puccinia sorghi fungus or both.
This invention also provides methods for manufacturing non-natural, transgenic seed that can be used to produce a crop of transgenic plants with an enhanced trait resulting from expression of a stably-integrated recombinant DNA construct. More specifically, the method comprises (a) screening a population of plants for an enhanced trait and a recombinant DNA construct, where individual plants in the population can exhibit the trait at a level less than, essentially the same as or greater than the level that the trait is exhibited in control plants, (b) selecting from the population one or more plants that exhibit the trait at a level greater than the level that said trait is exhibited in control plants, (c) collecting seed from a selected plant, (d) verifying that the recombinant DNA is stably integrated in said selected plants, (e) analyzing tissue of a selected plant to determine the production or suppression of a protein having the function of a protein encoded by nucleotides in a sequence of one of SEQ ID NOs: 1-3. In one aspect of the invention, the plants in the population further comprise DNA expressing a protein that provides tolerance to exposure to a herbicide applied at levels that are lethal to wild type plant cells and the selecting is affected by treating the population with the herbicide, e.g. a glyphosate, dicamba, or glufosinate compound. In another aspect of the invention the plants are selected by identifying plants with the enhanced trait. The methods are especially useful for manufacturing corn, soybean, cotton, canola, alfalfa, wheat, rice, sugarcane or sugar beet seed.
Another aspect of the invention provides a method of producing hybrid corn seed comprising acquiring hybrid corn seed from a herbicide tolerant corn plant which also has stably-integrated, recombinant DNA construct comprising a promoter that is (a) functional in plant cells and (b) is operably linked to DNA that encodes or suppresses a protein having the function of a protein encoded by nucleotides in a sequence of one of SEQ ID NOs: 1-3. The methods further comprise producing corn plants from said hybrid corn seed, wherein a fraction of the plants produced from said hybrid corn seed is homozygous for said recombinant DNA, a fraction of the plants produced from said hybrid corn seed is hemizygous for said recombinant DNA, and a fraction of the plants produced from said hybrid corn seed has none of said recombinant DNA; selecting corn plants which are homozygous and hemizygous for said recombinant DNA by treating with an herbicide; collecting seed from herbicide-treated-surviving corn plants and planting said seed to produce further progeny corn plants; repeating the selecting and collecting steps at least once to produce an inbred corn line; and crossing the inbred corn line with a second corn line to produce hybrid seed.
Another aspect of the invention provides a method of selecting a plant comprising plant cells of the invention by using an immunoreactive antibody to detect the presence or absence of protein expressed or suppressed by recombinant DNA in seed or plant tissue. Yet another aspect of the invention provides anti-counterfeit milled seed having, as an indication of origin, plant cells of this invention.
Still other aspects of this invention relate to transgenic plants with enhanced water use efficiency or enhanced nitrogen use efficiency. For instance, this invention provides methods of growing a corn, cotton, soybean, or canola crop without irrigation water comprising planting seed having plant cells of the invention which are selected for enhanced water use efficiency. Alternatively methods comprise applying reduced irrigation water, e.g. providing up to 300 millimeters of ground water during the production of a corn crop. This invention also provides methods of growing a corn, cotton, soybean or canola crop without added nitrogen fertilizer comprising planting seed having plant cells of the invention which are selected for enhanced nitrogen use efficiency.
Another aspect of the invention provides transgenic plants with enhanced oil levels, including algae. In a particular embodiment, the invention provides transgenic seeds with seed composition improvement including enhanced protein, oil or starch levels.
Some of the recombinant DNA provided by this invention are transcription factors. This invention provides a method of producing a transgenic plant having an enhanced agronomic trait produced by expression of a transcription factor. This method includes identifying target genes of a transcription factor, which includes the steps of assessing a dataset of expression profiles of a transcription factor gene and other genes and analyzing said dataset to determine a subset of genes that are regulated by said transcription factor, and cloning the coding sequence of at least one of the subset of genes into a plant transformation vector and transforming a plant with such vector.
Furthermore, this invention provides novel genes of Glycine max GLABRA2 (Gm.GL2) and Rhodosporidium toruloides DGAT2 (Rt.DGAT2); and an Arabidopsis/Brassica chimeric DNA construct (At.Bn.Otf1) of an oil transcription factor, coding for proteins as set forth in SEQ ID NO:4 through SEQ ID NO:6, respectively, which are particularly useful for generating transgenic crop plants having seeds with enhanced oil levels.