Modifications of transgenic corn event mon95275 and methods thereof

Abstract

Modified corn event MON95275 and plants, plant cells, seeds, plant parts, progeny plants, commodity products comprising detectable amounts of a modified corn event MON95275 DNA are provided. Polynucleotides and sequences specific for a modified corn event MON95275, methods for making a modified corn event MON95275, methods for making and using plants, plant cells, seeds, plant parts, progeny plants, and commodity products comprising a modified corn event MON95275, and methods for detecting a modified corn event MON95275, and polynucleotides or DNA sequences specific for a modified corn event MON95275, in a DNA molecule or sample, are provided.

Description

MODIFICATIONS OF TRANSGENIC CORN EVENT MON95275 AND METHODS THEREOFREFERENCE TO RELATED APPLICATION

[0001] This application claims the priority of U.S. Provisional Appl. Ser. 63 / 606,442, filed December 5, 2023, the entire disclosure of which is incorporated herein by reference.INCORPORATION OF SEQUENCE LISTING

[0002] The sequence listing contained in the file named BCS236358WO _ST26.xml is 311,478 bytes (measured in Microsoft Windows®), was created on September 11, 2024, is filed herewith by electronic submission, and is incorporated herein by reference.FIELD OF THE INVENTION

[0003] The present invention relates to recombinant DNA molecules present in and / or isolated from a modified corn event MON95275. The invention also relates to transgenic com plants, plant parts, seed, pollen, cells, and agricultural products containing a modified corn event MON95275, as well as methods of using the same, and making and detecting the presence of a modified corn event MON95275. Transgenic com plants, plant parts, seed and cells containing a modified corn event MON95275 DNA may exhibit resistance to infestations by insects in the family Coleoptera.BACKGROUND

[0004] Corn (zea mays) is an important crop and is a primary food source in many areas of the world. The methods of biotechnology have been applied to corn for improving agronomic traits and product quality. One such agronomic trait is insect resistance, manifested through the insertion of a recombinant DNA segment into the genome of the com plant.

[0005] There are a number of different transgenic events in com that have been described in the art that provide various types of insect resistance, particularly to Lepidopteran or Coleopteran species, and these include MON810, TC1507, MON89034, MON95379, and MIR162 among those that confer Lepidopteran resistance, and MON863, MON88017, DAS-59122-7, DP-004114-3, and DP23211 and MIR604 among those that confer Coleopteran resistance, particularly resistance to corn rootworm infestations. These transgenic events have been in use commercially in a variety of geographies across the glove for an extended period of time, often have used the same or similar toxins that were in use in earlier deployed transgenic events, and resistance to the expressed toxins in these events by targeted insect pests has been observed in many geographic regions where these have been deployed.

[0006] There is a continuing need in the art to provide novel transgenic events in corn that exhibit resistance to insect infestation, and preferably the novel transgenic events confer resistance to the target insects, including those races that have evolved resistance to the existing commercially deployed traits, using modes of action that are not overlapping with or similar to the modes of action previously deployed in earlier commercial embodiments. The present disclosure provides for modifications of a novel transgenic event that confers resistance to corn rootworm infestations, including resistance to rootworms that have evolved resistance to commercial embodiments that have been previously deployed.SUMMARY OF THE INVENTION

[0007] In one aspect, the present disclosure provides a recombinant DNA molecule comprising: (a) a first nucleotide sequence (i) comprising an expression cassette that encodes a Cry75Aal or Vip4Da2 protein or a DvSnf7 specific dsRNA, and / or (ii) selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7 and SEQ ID NO: 8; and (b) a second nucleotide sequence that (i) comprises at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 11 or 47, or (b) is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8% or at least 99.9% identical to at least 10, at least 1 1, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, atleast 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 1 1 or 47, or (iii) is selected from the group consisting of SEQ ID NOs: 49-148. In some embodiments, the first nucleotide sequence of the recombinant DNA molecule comprises at least two expression cassettes comprising a first expression cassette and a second expression cassette, wherein the first expression cassette encodes a Cry75Aal and the second expression cassette encodes a Vip4Da2 protein, or wherein the first expression cassette encodes a Cry75Aal and the second expression cassette encodes a DvSnf7 specific dsRNA, or wherein the first expression cassette encodes a Vip4Da2 and the second expression cassette encodes a DvSnf7 specific dsRNA. In some embodiments, the first nucleotide sequence of the recombinant DNA molecule comprises at least three expression cassettes comprising a first expression cassette, a second expression cassette and a third expression cassette, wherein the first expression cassette encodes a Cry75Aal, the second expression cassette encodes a Vip4Da2 protein, and the third expression cassette encodes a DvSnf7 specific dsRNA. In some embodiments, the recombinant DNA molecule may further comprise (c) a third nucleotide sequence that (i) comprises at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 12 or 48, or (ii) is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99. 1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8% or at least 99.9% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 12 or 48, or (iii) is selected from the group consisting of SEQ ID NOs: 149-248. The third nucleotide sequence, in certain embodiments, is selected from the group consisting of SEQ ID NOs: 149-248. The recombinant DNA molecule, in particular embodiments, has a deletion of one or more consecutivenucleotides between the second nucleotide sequence and the third nucleotide sequence relative to SEQ ID NO: 10, 12, or 48. In various embodiments, the second nucleotide sequence is selected from the group consisting of SEQ ID NOs: 49-148. In some embodiments, the recombinant DNA molecule has a deletion of one or more consecutive nucleotides between the first nucleotide sequence and the second nucleotide sequence relative to SEQ ID NO: 10, 11, or 47.

[0008] In another aspect, the present disclosure provides a recombinant DNA molecule comprising: (a) a first nucleotide sequence (i) comprising an expression cassette that encodes a Cry75Aal or Vip4Da2 protein or a DvSnf7 specific dsRNA, and / or (ii) selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, and SEQ ID NO: 9; and (b) a second nucleotide sequence that (i) comprises at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 12 or 48, or (ii) is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99. 1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8% or at least 99.9% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 12 or 48, or (iii) is selected from the group consisting of SEQ ID NOs: 149-248. In some embodiments, the first nucleotide sequence of the recombinant DNA molecule comprises at least two expression cassettes comprising a first expression cassette and a second expression cassette, wherein the first expression cassette encodes a Cry75Aal and the second expression cassette encodes a Vip4Da2 protein, or wherein the first expression cassette encodes a Cry75Aal and the second expression cassette encodes a DvSnf7 specific dsRNA, or wherein the first expression cassette encodes a Vip4Da2 and the second expression cassette encodes a DvSnf7 specific dsRNA. In some embodiments, the first nucleotide sequence of the recombinant DNA molecule comprises at leastthree expression cassettes comprising a first expression cassette, a second expression cassette and a third expression cassette, wherein the first expression cassette encodes a Cry75Aal, the second expression cassette encodes a Vip4Da2 protein, and the third expression cassette encodes a DvSnf7 specific dsRNA. In certain embodiment, the second nucleotide sequence is selected from the group consisting of SEQ ID NOs: 149-248. The recombinant DNA molecule, in some embodiments, has a deletion of one or more consecutive nucleotides between the first nucleotide sequence and the second nucleotide sequence relative to SEQ ID NO: 10, 12, or 48. In particular embodiments, the recombinant DNA molecule may further comprise nucleotides 1-1,073 or 15,756-16,861 of SEQ ID NO: 10.

[0009] In many embodiments, a recombinant DNA molecule of the present disclosure may be comprised in a corn plant, com plant part, corn plant cell, corn plant seed, corn progeny plant, or commodity or fuel product made from corn and com plant parts. In some embodiments, a recombinant DNA molecule of the present disclosure may comprise an amplicon diagnostic for the presence of DNA comprising a sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, and SEQ ID NO: 9. A recombinant DNA molecule of the present disclosure, in particular embodiments, is derived from a corn plant, corn plant part, corn seed, processed corn seed, corn plant cell or tissue, animal feed comprising com, corn oil, corn meal, com flour, corn flakes, corn bran, food made comprising com, com biomass, or fuel products made from corn and corn plant parts.

[0010] In yet another aspect, the present disclosure provides a recombinant DNA molecule comprising a polynucleotide segment of sufficient length to function as a DNA probe that hybridizes specifically under stringent hybridization conditions with a polynucleotide having a sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 47, and SEQ ID NO: 48. In still yet another aspect, the present disclosure provides a pair of DNA molecules that can function as DNA primers when used together in an amplification reaction comprising a first DNA molecule and a second DNA molecule, wherein the first DNA molecule and the second DNA molecule are different, (a) wherein the first DNA molecule is: (i) at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, or at least 99.9% identical or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, or SEQ ID NO: 9, or a complement thereof, or (ii) at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, or at least 99.9% identical or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 11 or 47, or a complement thereof; and (b) wherein the second DNA molecule is: (i) at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99. 1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, or at least 99.9% identical or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, or SEQ ID NO: 9, or a complement thereof, or (ii) at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, or at least 99.9% identical or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 11 or 47, or a complement thereof, or (ii) at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, or at least 99.9% identical or 100% identical to at least 10, at least 1 1, at least 12, at least 13, at least 14,at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21 , at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 11 or 47, or a complement thereof, or (iii) at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, or at least 99.9% identical or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 12 or 48, or a complement thereof. In certain embodiments, the first DNA molecule and the second DNA molecule can be used together in an amplification reaction with a sample from a plant, plant part, plant seed, plant cell, food or animal feed, or commodity or fuel product made from a plant or plant part to produce an amplicon diagnostic for the presence of a modified com event MON95275 DNA in said sample. The amplicon, in some embodiments, comprises a nucleotide sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10.

[0011] In one aspect, the present disclosure provides a method of detecting the presence of a DNA segment diagnostic for a modified com event MON95275 DNA in a sample, said method comprising: contacting the sample with the DNA molecule that functions as a DNA probe of the present disclosure; subjecting the sample and the DNA molecule to stringent hybridization conditions; and detecting hybridization of the DNA molecule to the DNA segment in the sample, wherein the detection is diagnostic for the presence of the modified corn event MON95275 DNA in the sample. In another aspect, the present disclosure provides a method of detecting the presence of a DNA segment diagnostic for a modified com event MON95275 DNA in a sample, the method comprising: contacting the sample with a pair of DNA molecules capable of functioning as DNA primers when used together in an amplification reaction provided by the present disclosure; performing an amplification reaction sufficient to produce a DNA amplicon; and detecting the presence of the DNA amplicon in the reaction, wherein the presence of the DNA amplicon is diagnostic for the presence of the modified com event MON95275 DNA in the sample. In yet another aspect, the present disclosure provides a method of detecting the presence of a DNAsegment diagnostic for a modified corn event MON95275 DNA in a sample, the method comprising performing a sequencing reaction with the sample, wherein the production in the sequencing reaction of a target nucleotide sequence comprising at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, or SEQ ID NO: 9, or a complement thereof, is diagnostic for the modified corn event MON95275 DNA in the sample. In particular embodiments, the modified corn event MON95275 is a further modified corn event MON95275. In yet another embodiment, the present disclosure provides a DNA detection kit comprising: (a) a DNA probe of the present disclosure; and / or (b) a pair of DNA molecules that function as DNA primers as provided by the present disclosure.

[0012] In another aspect, the present disclosure provides a modified corn plant, com plant part, corn seed, or com cell comprising a modified com event MON95275 or comprising a recombinant DNA molecule as provided herein. In some embodiments, the modified com plant, corn plant part, corn seed, or corn cell may comprise a recombinant DNA molecule or DNA segment comprising a nucleotide sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10, or a complement thereof; or a or a recombinant DNA molecule or DNA segment comprising a nucleotide sequence that is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 47, or SEQ ID NO: 48, or a complement thereof. According to present embodiments, the corn plant, corn plant part, corn seed, or corn cell exhibitsresistance to Western Com Rootworm (Diahrotica virgifera virgifera, WCR) and / or Northern Corn Rootworm Diabrotica barberi, NCR). In particular embodiments, the corn plant, com plant part, corn seed, or corn cell is further defined as a progeny plant of any generation of a corn plant comprising a modified corn event MON95275, or a com plant part, corn seed, or corn cell derived therefrom. In some embodiments, a recombinant DNA molecule comprises all or part of chromosome 3 of a com or maize genome, or a DNA segment is present in chromosome 3 of a modified com plant, corn plant part, corn seed, or corn cell. According to some embodiments, a modified corn event MON95275 of a modified com plant, corn plant part, corn seed, or com cell comprises a genetic modification, mutation or edit, relative to the corn event MON95275, introduced via a mutagenesis or targeted genome editing technique.

[0013] In yet another aspect, the present disclosure provides a method of producing a progeny corn plant comprising a modified com event MON95275 comprising: (a) sexually crossing a first modified com plant that comprises a modified corn event MON95275 with itself or a second corn plant; (b) collecting one or more seeds produced from said cross; (c) growing said seed to produce one or more progeny plants; and (d) selecting at least a first progeny plant or seed comprising a modified com event MON95275. The present disclosure, in additional embodiments, provides a hybrid modified corn plant or seed comprising a modified com event MON95275 produced by the methods described herein. In a further embodiment, the method described herein further comprises: (e) collecting seed from said at least first progeny plant comprising a modified corn event MON95275. In some embodiments, the hybrid modified corn plan or seed of claim 39, wherein the modified com event MON95275 is a further modified corn event MON95275.

[0014] Aspects of the present disclosure provide a nonliving com plant material comprising a detectable amount of a recombinant DNA molecule as described herein. In some embodiments, the present disclosure provides microorganisms and commodity products comprising a recombinant DNA molecule of the present disclosure. The microorganism, in a number of embodiments, is a plant cell. In some embodiments, a commodity product is produced from a modified com plant, corn plant part, corn seed, or corn tissue or cell comprising a modified corn event MON95275. The modified corn event MON95275, in certain embodiments, is a further modified corn event MON95275. Non-limiting examples of commodity products include whole or processed com seed, animal feed comprising corn, corn oil, com meal, corn flour, corn flakes, corn bran, corn biomass, and fuel products produced using com and com plant parts. In certainaspects, the present disclosure provides a method of producing a commodity product, the method comprising: (a) obtaining a modified corn plant, corn plant part, or corn seed comprising a modified com event MON95275; and (b) producing a commodity product from the transgenic corn plant, corn plant part, or com seed.

[0015] In aspects of the present disclosure, a modified corn event MON95275 may be a further modified corn event MON95275.

[0016] In one aspect, the present disclosure provides a com plant, corn plant part, or corn seed comprising a DNA molecule or segment functional as a template when tested in a DNA amplification method to produce an amplicon diagnostic for the presence of a modified corn event MON95275 DNA.

[0017] In another aspect, the present disclosure provides a method of determining the zygosity of a corn plant, corn plant part, or corn seed comprising a modified corn event MON95275 comprising: (a) contacting a sample comprising DNA from the com plant, corn plant part, or corn seed with a pair of DNA molecules that function as DNA primers, as provided by the present disclosure; performing a nucleic acid amplification reaction with the sample and the pair of DNA molecules; and detecting in the nucleic acid amplification reaction a first amplicon diagnostic for a modified corn event MON95275 and a second amplicon diagnostic for native corn genomic DNA not comprising the modified com event MON95275, wherein the presence of only the first amplicon is diagnostic of a com plant, com plant part, or corn seed homozygous for the modified corn event MON95275, and the presence of both the first amplicon and the second amplicon is diagnostic of a corn plant, com plant part, or com seed heterozygous for the modified corn event MON95275. In yet another aspect, the present disclosure provides a method of determining the zygosity of a com plant, com plant part, or corn seed comprising a modified com event MON95275 and a second primer pair that can produce a second amplicon of a standard genomic sequence known to be single copy and homozygous in the com plant, com plant part, or corn seed; contacting the sample with a first probe that specifically hybridizes to the first amplicon and / or all or part of the modified com event MON95275, performing a DNA amplification reaction using real-time PCR with the sample and determining the cycle thresholds (Ct values) of the first amplicon and the second amplicon; calculating the difference (ACt) between the Ct values of the second amplicon and the first amplicon; and determining the zygosity of the modified corn event MON95275, wherein a ACt of about zero (0) indicates homozygosity of the modified corn eventMON95275 and a ACt of about one (1) indicates heterozygosity of the modified com event MON95275. In some embodiments, the first and second primer pairs comprise SEQ ID NO: 15 combined with SEQ ID NO: 16, and SEQ ID NO: 18 combined with SEQ ID NO: 19.

[0018] In still yet another aspect, the present disclosure provides a method of determining the zygosity of a com plant, com plant part, or corn seed comprising a modified com event MON95275 comprising: contacting a sample comprising DNA from the com plant, corn plant part, or corn seed with a primer pair capable of producing a first amplicon diagnostic for the modified corn event MON95275 and a second amplicon diagnostic for native corn genomic DNA not comprising the modified com event MON95275; performing a nucleic acid amplification reaction with the sample and the set of primer pairs; and detecting the first amplicon and the second amplicon, wherein the presence of only the first amplicon is diagnostic of a com plant, corn plant part, or corn seed homozygous for the modified corn event MON95275, the presence of only the second amplicon is diagnostic of a corn plant, corn plant part, or com seed homozygous for native com genomic DNA not comprising the modified com event MON95275, and the presence of both the first amplicon and the second amplicon is diagnostic of a corn plant, corn plant part, or com seed heterozygous for the modified corn event MON95275. In one aspect, the present disclosure provides a method of determining the zygosity of a corn plant, corn plant part, or corn seed comprising a modified corn event MON95275 comprising: contacting a sample comprising DNA from the corn plant, corn plant part, or corn seed with a probe set which contains at least a first probe that specifically hybridizes to the modified corn event MON95275 and at least a second probe that specifically hybridizes to corn genomic DNA that was disrupted by insertion of the heterologous DNA of com event MON95275 and is disrupted by the modified corn event MON95275, wherein the second probe does not hybridize to the modified corn event MON95275 DNA; and hybridizing the probe set with the sample under stringent hybridization conditions, wherein detecting hybridization of only the first probe under the hybridization conditions is diagnostic for a corn plant, corn plant part, or com seed homozygous for the modified corn event MON95275. The probe set, in particular embodiments, comprises SEQ ID NO: 17 and SEQ ID NO: 20. In some embodiments, the modified com event MON95275 is a further modified corn event MON95275.

[0019] Aspects of the present disclosure provide a population of transgenic corn plants, wherein each transgenic corn plant comprises a modified com event MON95275. In some embodiments,the population of com plants have an increased resistance to Coleopteran insect pest species on average relative to a population of control corn plants lacking the modified corn event MON95275. The modified com event MON95275, in additional embodiments, is a further modified com event MON95275. In particular embodiments, the population of com plants has an increased resistance to a Lepidopteran insect pest species on average relative to a population of control com plants lacking the modified com event MON95275.

[0020] In some aspect, the present disclosure provides a method of modifying a corn plant, the method comprising: (a) introducing a site-specific nuclease or a recombinant DNA construct comprising an expression cassette encoding a site-specific nuclease into at least one cell of an explant of a corn plant comprising corn event MON95275, or a plant part thereof, to produce a modified corn event MON95275 via a targeted genome editing technique; and (b) developing or regenerating a modified corn plant from the explant, wherein the modified corn plant comprises the modified corn event MON95275. The modified corn event MON95275, in additional embodiments, is a further modified corn event MON95275. Non-limiting examples site-specific nucleases include a zinc-finger nuclease (ZFN), a meganuclease, an RNA-guided endonuclease, a TALE-endonuclease (TALEN), a recombinase, and a transposase. In some embodiments, the sitespecific nuclease is an RNA-guided endonuclease or a CRISPR / Cas nuclease. The introducing step (a), in particular embodiments, comprises introducing the recombinant DNA construct into the at least one cell of the explant, wherein the recombinant DNA construct further comprises an expression cassette encoding a first guide RNA (gRNA). The recombinant DNA construct, in additional embodiments, further comprises an expression cassette encoding a second guide RNA (gRNA). The introducing step (a), in certain embodiments, further comprises introducing a first guide RNA (gRNA) or a second recombinant DNA construct comprising an expression cassette encoding a first guide RNA (gRNA) into the at least one cell of the explant. The introducing step (a), in some embodiments, comprises introducing at least two guide RNAs (gRNAs) comprising the first gRNA and a second gRNA into the at least one cell of the explant. In a number of embodiments, the second recombinant DNA construct further comprises an expression cassette encoding a second guide RNA (gRNA). The introducing step (a), in many embodiments, comprises introducing a first gRNA and second recombinant DNA construct comprising an expression cassette encoding a second guide RNA (gRNA) into the at least one cell of the explant. In some embodiments, the site-specific nuclease has a first target site in the genome of the complant at or near corn event MON95275. In particular embodiments, the site-specific nuclease has a second target site in the genome of the com plant at or near com event MON95275. The introducing step (a), in certain embodiments, comprises introducing a second site-specific nuclease or a recombinant DNA construct comprising an expression cassette encoding a second site-specific nuclease into at least one cell of the explant, and wherein the second site-specific nuclease has a second target site in the genome of the corn plant at or near corn event MON95275. In some embodiments, the first gRNA has a first target site in a flanking DNA sequence, 5 ' flank, 3 ' flank, junction sequence, or insertion sequence of com event MON95275, or a complement thereof. In particular embodiments, the first gRNA has a first target site comprising a target sequence that is: (i) at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 11 or 47, or a complement thereof; or (ii) at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides SEQ ID NO: 12 or 48, or a complement thereof;; or (iii) at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, or SEQ ID NO: 9, or a complement thereof. In some embodiments, the first gRNA has a second target site comprising a target sequence that is: (i) at least 90%, at least 91%, at least 92%, at least 93%, atleast 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 1 1, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 11 or 47, or a complement thereof; or (ii) at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 12 or 48, or a complement thereof; or (iii) at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99. 1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, or SEQ ID NO: 9, or a complement thereof. In certain embodiments, (i) the first gRNA has a first target site comprising a target sequence that is: (1) at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 11 or 47, or a complement thereof; or (2) at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99. 1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least17, at least 18, at least 19, at least 20, at least 21 , at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 12 or 48, or a complement thereof; or (3) at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 1 1, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, or SEQ ID NO: 9, or a complement thereof; and (ii) the second gRNA has a second target site comprising a target sequence that is: (ii) wherein the second gRNA has a second target site comprising a target sequence that is: (1) at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 11 or 47, or a complement thereof; or (2) at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99. 1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 1 1, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 12 or 48, or a complement thereof; or (3) at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO:4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, or SEQ ID NO: 9, or a complement thereof. In some embodiments, the modified corn event MON95275 comprises a deletion or excision of intervening genomic DNA between the first target site and the second target site, relative to the corn event MON95275. In certain embodiments, the methods of the present disclosure may further comprise selecting the modified com plant comprising the modified corn event MON95275, and sexually crossing the modified com plant with itself or a second corn plant to produce one or more modified progeny corn plants.

[0021] Aspect of the present disclosure provide a method of introducing a target site into a com plant, the method comprising: (a) introducing a cognate target site into the com event MON95275 locus of at least one cell of a corn plant or corn plant part comprising the corn event MON95275 or an explant thereof via a targeted genome editing technique, wherein the cognate target site is identical or similar to an originator target site for a site-specific nuclease present in the com event MON95275 locus, and (b) developing or regenerating a modified corn plant comprising a modified corn event MON95275 comprising the cognate target site. In some embodiments, the methods of the present disclosure may further comprise: (c) introducing a site-specific nuclease or a recombinant DNA construct comprising an expression cassette encoding a site-specific nuclease into at least one cell of an explant of a corn plant comprising the modified corn event MON95275 or a plant part thereof, to produce a further modified corn event MON95275 via a targeted genome editing technique, wherein the target site of the site-specific nuclease includes the cognate target site and the originator target site; and (d) developing or regenerating a second modified corn plant comprising the further modified com event MON95275.

[0022] In certain aspects, the present disclosure provides a method of introducing a target site into a corn plant, the method comprising: (a) introducing a cognate target site into the corn event MON95275 locus of at least one cell of a corn plant or corn plant part comprising the corn event MON95275 or an explant thereof via a targeted genome editing technique to produce a modified corn event MON95275 comprising the cognate target site, wherein the cognate target site is identical or similar to an originator target site for a site-specific nuclease present in the com event MON95275 locus, and (b) introducing a site-specific nuclease or a recombinant DNA construct comprising an expression cassette encoding a site-specific nuclease into at least one cell of an explant of a modified corn plant comprising the modified com event MON95275, or a plant part thereof, to produce a further modified com event MON95275 via a targeted genome editingtechnique, wherein the target site of the site-specific nuclease includes the cognate target site and the originator target site; and (c) developing or regenerating a second modified corn plant comprising the further modified corn event MON95275. In some embodiments, the further modified corn event MON95275 of the second modified corn plant comprises a deletion or excision of intervening genomic DNA between the originator target site and the cognate target site, relative to the corn event MON95275 or modified corn event MON95275. In certain embodiments, the methods of the present disclosure may further comprise selecting the second modified corn plant or a progeny plant of the second modified corn plant comprising the further modified corn event MON95275, and sexually crossing the second modified corn plant or the progeny plant with itself or another corn plant to produce one or more modified progeny corn plants comprising the further modified corn event MON95275. In some embodiments, the modified corn event MON95275 is a further modified corn event MON95275.

[0023] In some aspects, the present disclosure provides a method of modifying an explant of a corn plant or plant part, the method comprising: introducing a site-specific nuclease or a recombinant DNA construct comprising an expression cassette encoding a site-specific nuclease into at least one cell of an explant of a corn plant or plant part comprising corn event MON95275 to produce a modified corn event MON95275 into the at least one cell of the explant. In certain embodiments, the modified corn event MON95275 is a further modified com event MON95275.BRIEF DESCRIPTION OF THE DRAWINGS

[0024] FIG. 1 is a graphical depiction of the orientation and alignment of the DNA elements / segments that are present within the nucleotide sequence shown in SEQ ID NO: 10, which is the sequence of the inserted transgenic DNA and the corresponding adjacent 5' and 3' sequences of the corn genome present within the corn event MON95275. [1] (SEQ ID NO: 1) and [2] (SEQ ID NO: 2) each graphically represent the approximate positions of the sequences of the 50 consecutive nucleotide segments referred to respectively as a 5' or 3' junction sequence, respectively the arbitrarily assigned left, 5 ' end, and the right, 3 ' end junction sequences of [9] that consist respectively of 25 consecutive nucleotides of corn genome DNA (ends of lighter gray shaded segment of

[0010] ) and 25 consecutive nucleotides of adjacent inserted transgenic DNA (darker gray shaded segment of

[0010] ); [3] (SEQ ID NO: 3) and [4] (SEQ ID NO: 4) each graphically represent 100 consecutive nucleotide segments of DNA at the 5' and 3' junctionpositions, are respectively a 5' or 3' junction sequence, and each contain 50 consecutive nucleotides of com genome DNA and 50 consecutive nucleotides of adjacent inserted transgenic DNA; [5] (SEQ ID NO: 5) and [6] (SEQ ID NO: 6) each graphically represent 200 consecutive nucleotide segments of DNA at the junction positions, are respectively a 5 ' or 3 'junction sequence, and each contain 100 consecutive nucleotides of corn genome DNA and 100 consecutive nucleotides of adjacent inserted transgenic DNA; [7] (SEQ ID NO: 7) is representative of the 5' junction region of corn genomic DNA and the inserted transgenic DNA and contains 1,073 consecutive nucleotides of the corn genome DNA and 153 consecutive nucleotides of the adjacent inserted transgenic DNA; [8] (SEQ ID NO: 8) is representative of the 3' junction region of corn genomic DNA and the inserted transgenic DNA containing 101 consecutive nucleotides of the inserted transgenic DNA and 1,006 consecutive nucleotides of the adjacent corn genome DNA; [9] (SEQ ID NO: 9) represents the length and structure of the inserted DNA, and the arrows and labels below each arrow represent the expression elements in the three cassettes within the inserted DNA in which RB / LB represent the positions of the right and left borders of the Agrobacterium double border mediated transformation vector, LoxP represents the position of the residual Cre- recombinase recognition site remaining in the inserted DNA after marker excision, the three letter E’s represent the positions of enhancer elements in the respective constructs, the three letter P’s represent the positions of the promoter elements in the respective constructs, the three letter L’s represent the positions of leader sequences (5 ' untranslated regions, 5 UTR) in the respective constructs, the three letter l’s represent the positions of the intron sequences in the respective constructs, the three letter T’s represent the positions of the transcription termination sequences (3 ' untranslated regions, 3 UTR) in the respective constructs, and ISR represents the position of an intergenic sequence region (ISR4). The three constructs from right to left on the page of the drawing encode the coleopteran pest toxic Vip4Da2 and Cry75Aal toxins, and the segment encoding an RNA molecule capable of folding into a hairpin shaped double stranded molecule that is designed for suppression of transcripts from and thus reduction of the translated protein, Snf7, a protein that is essential for survival of corn rootworm larvae.

[0011] (SEQ ID NO: 11) is representative of the position of the com genome DNA flanking the 5' end of the inserted DNA, and

[0012] (SEQ ID NO: 12) is representative of the position of the corn genome DNA flanking the 3 ' end of the inserted DNA.

[0015] (SEQ ID NO: 15, primer SQ51355), and

[0016] (SEQ ID NO: 16, primer SQ51355) are representative of the position of a primer pair that can be used in a thermalamplification reaction to produce an amplicon of 74 nucleotides containing the right insert / genome junction, the arrows showing the direction in which the amplification would proceed to form the amplicon from the respective positions within

[0010] ,

[0017] (SEQ ID NO: 17, PB10263) is representative of a probe and the position to which the probe would bind (or hybridize to) the amplicon produced using primers

[0016] and

[0017] , for detecting the presence of the MON95275 Event in a sample.

[0025] FIG. 2 illustrates the T-DNA cassette in the plasmid vector used to transform corn. One insertion event, when subjected to Cre-recombinase marker excision, resulted in event MON95275.

[0013] (SEQ ID NO: 13) illustrates the DNA in the plasmid vector before integration (the “T-DNA Before Integration”). The arrows below

[0013] represent the individual genetic elements comprised within the three transgene cassettes designed to express the result effective coleopteran toxic agents. The CP4 EPSPS selectable marker cassette is flanked between the two LoxP segments which are recognized by the Cre-recombinase and which is capable of excising the selectable marker from the insertion event containing

[0013] , The insertion event DNA is represented by

[0014] , differing from

[0013] only by the fact that the segment

[0013] has been inserted into the corn genome, and is now depicted as being flanked 5 ' and 3 ' by the corn genome segments labeled as 5' Flank and 3' Flank.

[0018] represents the segment shown in Figure 1 as

[0010] ,

[0026] Figure 3 is a diagrammatic representation of the timeline for the research, testing, and development that was relied upon in order to select the corn event MON95275.BRIEF DESCRIPTION OF THE SEQUENCES

[0027] SEQ ID NO: 1 is a 50-nucleotide sequence representing the 5 ' junction region of corn genomic DNA and the integrated transgenic expression cassette (25 nucleotides corn genome DNA at 5' end of SEQ ID NO: 1, 25 nucleotides transgenic inserted DNA at 3' end of SEQ ID NO: 1) , and can be identified within SEQ ID NO: 10 at nucleotide positions 1,049-1,098.

[0028] SEQ ID NO: 2 is a 50-nucleotide sequence representing the 3' junction region of the integrated transgenic expression cassette and the com genomic DNA (25 nucleotides transgenic inserted DNA at 5' end of SEQ ID NO: 2, 25 nucleotides corn genome DNA at 3' end of SEQ ID NO: 2), and can be identified within SEQ ID NO: 10 at nucleotide positions 15,731-15,780.

[0029] SEQ ID NO: 3 is a 100-nucleotide sequence representing the 5' junction region of corn genomic DNA and the integrated transgenic expression cassette (50 nucleotides corn genomeDNA at 5' end of SEQ ID NO: 3, 50 nucleotides transgenic inserted DNA at 3' end of SEQ ID NO: 3), and can be identified within SEQ ID NO: 10 at nucleotide positions 1,024-1,123.

[0030] SEQ ID NO: 4 is a 100-nucleotide sequence representing the 3' junction region of the integrated transgenic expression cassette and the corn genomic DNA (50 nucleotides transgenic inserted DNA at 5 ' end of SEQ ID NO: 4, 50 nucleotides corn genome DNA at 3 ' end of SEQ ID NO: 4), and can be identified within SEQ ID NO: 10 at nucleotide positions 15,706-15,805.

[0031] SEQ ID NO: 5 is a 200-nucleotide sequence representing the 5' junction region of corn genomic DNA and the integrated transgenic expression cassette (100 nucleotides corn genome DNA at 5' end of SEQ ID NO: 5, 100 nucleotides transgenic inserted DNA at 3' end of SEQ ID NO: 5) , and can be identified within SEQ ID NO: 10 at nucleotide positions 974-1,173.

[0032] SEQ ID NO: 6 is a 200-nucleotide sequence representing the 3' junction region of the integrated transgenic expression cassette and the com genomic DNA (100 nucleotides transgenic inserted DNA at 5' end of SEQ ID NO: 6, 100 nucleotides com genome DNA at 3' end of SEQ ID NO: 6), and can be identified within SEQ ID NO: 10 at nucleotide positions 15,656-15,855.

[0033] SEQ ID NO: 7 is a 1,226-nucleotide sequence representing the 5' junction region of corn genomic DNA and the integrated transgenic expression cassette (1,073 nucleotides com genome DNA at 5' end of SEQ ID NO: 5, 153 nucleotides transgenic inserted DNA at 3' end of SEQ ID NO: 5) , and can be identified within SEQ ID NO: 10 at nucleotide positions 1-1,226.

[0034] SEQ ID NO: 8 is a 1,207-nucleotide sequence representing the 3' junction region of the integrated transgenic expression cassette and the com genomic DNA (101 nucleotides transgenic inserted DNA at 5' end of SEQ ID NO: 8, 1,106 nucleotides com genome DNA at 3' end of SEQ ID NO: 8), and can be identified within SEQ ID NO: 10 at nucleotide positions 15,655-16,861.

[0035] SEQ ID NO: 9 is a 14,682-nucleotide sequence corresponding to the transgenic inserted T- DNA of corn event MON95275, and can be identified within SEQ ID NO: 10 at nucleotide positions 1,074-15,755.

[0036] SEQ ID NO: 10 is a 16, 861 -nucleotide sequence corresponding to the contiguous nucleotide sequence of the 5' genomic flanking DNA nucleotide sequence, the inserted T-DNA nucleotide sequence in event MON95275, and the 3 ' genomic flanking DNA nucleotide sequence; and includes SEQ ID NO: 11 (nucleotides 1-1,073), SEQ ID NO: 9 (nucleotides 1,074-15,755), and SEQ ID NO: 12 (nucleotides 15,756-16,861).

[0037] SEQ ID NO: 1 1 is a 1,073 -nucleotide sequence representing the com genomic DNA flanking the 5' end of the inserted T-DNA, and can be identified within SEQ ID NO: 10 at nucleotide positions 1-1,073.

[0038] SEQ ID NO: 12 is a 1,106-nucleotide sequence representing the com genomic DNA flanking the 3 ' end of the inserted T-DNA, and can be identified within SEQ ID NO: 10 at nucleotide positions 15,756-16,861.

[0039] SEQ ID NO: 13 is a 19,612-nucleotide sequence representing the transgene cassette comprised within the binary plant transformation plasmid vector used to transform corn to produce corn event MON95275.

[0040] SEQ ID NO: 14 is a 35-nucleotide LoxP sequence representing used for Cre-mediated excision and recombination, and the residual sequence can be identified within SEQ ID NO: 10 at nucleotide positions 15,444-15,478.

[0041] SEQ ID NO: 15 is a 27-nucleotide sequence corresponding to a thermal amplification primer referred to as SQ20267 which can be used to identify corn event MON95275 DNA in a sample or which can be used to detect an insertion event that, when subjected to Cre-recombinase marker excision, results in event MON95275 DNA. SEQ ID NO: 15 is identical to the nucleotide sequence corresponding to positions 15,706-15,732 of SEQ ID NO: 10.

[0042] SEQ ID NO: 16 is a 24-nucleotide sequence corresponding to a thermal amplification primer referred to as SQ51355 used to identify corn event MON95275 DNA in a sample or which can be used to detect an insertion event that, when subjected to Cre-recombinase marker excision, results in event MON95275 DNA. SEQ ID NO: 16 is identical to the reverse compliment of the nucleotide sequence corresponding to positions 15,756-15,779 of SEQ ID NO: 10.

[0043] SEQ ID NO: 17 is a 19-nucleotide sequence corresponding to a probe referred to as PB 10263 used to identify corn event MON95275 DNA in a sample or which can be used to detect an insertion event that, when subjected to Cre-recombinase marker excision, results in event MON95275 DNA. SEQ ID NO: 17 is identical to the nucleotide sequence corresponding to positions 15,734-15,752 of SEQ ID NO: 10, and as a probe can bind to a polynucleotide segment having the reverse complement of the nucleotides at this position.

[0044] SEQ ID NO: 18 is a 24-nucleotide sequence corresponding to a thermal amplification primer referred to as SQ20222 used as an internal control for the event and zygosity assay for corn event MON95275 and hybridizes to a region of the com genome.

[0045] SEQ ID NO: 19 is a 28-nucleotide sequence corresponding to a thermal amplification primer referred to as SQ20221 used as an internal control for the event and zygosity assay for corn event MON95275 and hybridizes to a region of the com genome.

[0046] SEQ ID NO: 20 is a 17-nucleotide sequence corresponding to a probe referred to as PB50298 used as an internal control for the event and zygosity assay for corn event MON95275 and hybridizes to a region of the com genome.

[0047] SEQ ID NO: 21 is a 20-nucleotide sequence corresponding to a thermal amplification primer referred to as PNEG95275_F used in the zygosity assay for corn event MON95275 and hybridizes to a region of corn genomic DNA which was deleted when the T-DNA used to produce event MON95275 inserted into the com genome. An amplicon produced in a thermal amplification reaction using the combination of primers PNEG95275 F and PNEG95275 R (SEQ ID NO: 22) and native corn DNA as template is diagnostic for the wild-type allele lacking the MON95275 inserted T-DNA.

[0048] SEQ ID NO: 22 is a 20-nucleotide sequence corresponding to a thermal amplification primer referred to as PNEG95275_R used in the zygosity assay for corn event MON95275 and hybridizes to a region of corn genomic DNA which was deleted when the T-DNA used to produce event MON95275 inserted into the com genome. An amplicon produced in a thermal amplification reaction using the combination of primers PNEG95275 F (SEQ ID NO: 21) and PNEG95275_R and native com DNA as template is diagnostic for the wild-type allele lacking the MON95275 inserted T-DNA.

[0049] SEQ ID NO: 23 is a 17-nucleotide sequence corresponding to a probe referred to as PRBNEG95275 used in the zygosity assay for confirming the absence of corn event MON95275 and hybridizes to a region of native corn genomic DNA which was deleted when the T-DNA used to produce event MON95275 inserted into the corn genome.

[0050] SEQ ID NO: 24 is a DNA sequence that functions in plants as an expression enhancer segment.

[0051] SEQ ID NO: 25 is a plant functional promoter operably linked to an untranslated leader sequence.

[0052] SEQ ID NO: 26 is a DNA sequence that functions in plants as an expression enhancer segment.

[0053] SEQ ID NO: 27 is a 27-nucleotide sequence corresponding to an OgRRS, OgRRS_5-l.

[0054] SEQ ID NO: 28 is a 27-nucleotide sequence corresponding to an OgRRS, OgRRS_5-2.

[0055] SEQ ID NO: 29 is a 27-nucleotide sequence corresponding to an OgRRS, OgRRS_In-l.

[0056] SEQ ID NO: 30 is a 27-nucleotide sequence corresponding to an OgRRS, OgRRS In-2.

[0057] SEQ ID NO: 31 is a 27-nucleotide sequence corresponding to an OgRRS, OgRRS_3-l.

[0058] SEQ ID NO: 32 is a 27-nucleotide sequence corresponding to an OgRRS, OgRRS_3-l.

[0059] SEQ ID NO: 33 is a 51-nucleotide sequence corresponding to a gRNA, gRNA_OgRRS_5- 1 comprised of a Casl2a protospacer adjacent motif (PAM) site operably linked to a guide-RNA hybridization site.

[0060] SEQ ID NO: 34 is a 51-nucleotide sequence corresponding to a gRNA, gRNA_OgRRS_5- 2.

[0061] SEQ ID NO: 35 is a 51-nucleotide sequence corresponding to a gRNA, gRNA OgRRS In- 1.

[0062] SEQ ID NO: 36 is a 51-nucleotide sequence corresponding to a gRNA, gRNA OgRRS In- 2.

[0063] SEQ ID NO: 37 is a 51-nucleotide sequence corresponding to a gRNA, gRNA_OgRRS_3- 1.

[0064] SEQ ID NO: 38 is a 51-nucleotide sequence corresponding to a gRNA, gRNA_OgRRS_3- 2.

[0065] SEQ ID NO: 39 is a sequence of a synthetic DNA coding sequence designed for expression in a plant cell encoding a nuclear targeted LbCasl2a CRISPR-associated protein.

[0066] SEQ ID NO: 40 is an amino acid sequence of a nuclear targeted LbCasl2a CRISPR- associated protein encoded by SEQ ID NO: 39.

[0067] SEQ ID NO: 41 is a sequence of a synthetic DNA coding sequence designed for expression in a plant cell encoding a nuclear targeted LbCasl2a-TYCV CRISPR-associated protein.

[0068] SEQ ID NO: 42 is an amino acid sequence of a nuclear targeted LbCasl2a-TYCV CRISPR-associated protein encoded by SEQ ID NO: 41.

[0069] SEQ ID NO: 43 is a sequence of a synthetic DNA coding sequence designed for expression in a plant cell encoding a nuclear targeted LbCasl2a-TATV CRISPR-associated protein.

[0070] SEQ ID NO: 44 is an amino acid sequence of a nuclear targeted LbCasl2a-TATV CRISPR-associated protein encoded by SEQ ID NO: 43.

[0071] SEQ ID NO: 45 is a sequence of a synthetic DNA coding sequence designed for expression in a plant cell encoding a nuclear targeted FnCasl2a CRISPR-associated protein.

[0072] SEQ ID NO: 46 is an amino acid sequence of a nuclear targeted FnCasl2a CRISPR- associated protein encoded by SEQ ID NO: 45.

[0073] SEQ ID NO: 47 is a 5000-nucleotide sequence representing corn genomic DNA that flanks the transgenic insert at the 5 end of the insert. Nucleotides 3,928-5,000 of SEQ ID NO: 47 are identical to nucleotides 1-1,073 of SEQ ID NO: 11. Nucleotides 1-3,927 are based on the genomic sequence of the B73 com cultivar.

[0074] SEQ ID NO: 48 is a 5,000-nucleotide sequence representing com genomic DNA that flanks the transgenic insert at the 3 ' end of the insert. Nucleotides 1-1,106 of SEQ ID NO: 48 are identical to nucleotides 1-1,106 of SEQ ID NO: 12. The remaining nucleotides (1,107-5,000) are based on the genomic sequence of the B73 com cultivar.

[0075] SEQ ID NOs:49-148 are 50-nucleotide sequence in the 5' flank genomic sequence of corn event MON95275.

[0076] SEQ ID NOs: 149-248 are 50-nucleotide sequence in the 3 ' flank genomic sequence of corn event MON95275.DETAILED DESCRIPTION

[0077] The present disclosure relates to a transgenic com event - MON95275 - that achieves insecticidal control over Coleopteran pests of corn by expression of Cry75Aal, Vip4Da2, and a dsRNA targeting for suppression the native and essential corn rootworm DvSnf7. Specifically, corn event MON95275 provides resistance to the Coleopteran insect pests Western Corn Rootworm (Diabrotica virgifera virgifera, WCR) and Northern Corn Rootworm (Diabrotica barberi, NCR). Corn event MON95275 will meet a great need for control of these insects in corn agricultural production where corn rootworms are present, because chemical insecticides often do not provide adequate control of these insects, or because multiple applications of such chemistries are required to be applied throughout the growing season, increasing the labor requirements, carbon footprint, and input of chemical pesticides in the environment as well as adding significantly to the cost of corn production. Reference to corn event MON95275 herein is intended as being equivalent to a reference to MON 95275, event MON95275, event MON 95275, MON95275 event, MON 95275 event; the references are interchangeable.

[0078] The resistance to infestation by Coleopteran species provided by event MON95275 arises in connection with the expression of a DNA segment encoding two insecticidal proteins and a double-stranded RNA (dsRNA) capable of interfering with a corn root worm essential gene, that are operably and covalently linked within the inserted transgenic DNA that in part defines the corn event MON95275. The two insecticidal proteins in the MON95275 event are a Cry75Aal protein (United States Patent Application Publication No. 2016-0319302A2, SEQ ID NO: 25, coding sequence, SEQ ID NO: 37) and a Vip4Da2 protein (United States Patent No. 10,100,330, SEQ ID NO: 2, coding sequence, SEQ ID NO: 3). The dsRNA produced in the event MON95275 targets for suppression a gene referred to as DvSnf7, in Western Corn Rootworm (Diabrotica virgifera virgifera when ingested by a rootworm (see, for example, United States Patent No. 7,943,818, SEQ ID NO: 818). These two insecticidal proteins and dsRNA are expressed from the three expression cassettes within the inserted transgenic DNA construct as set forth in SEQ ID NO: 9 and illustrated in Fig. 1.

[0079] The Ciy75Aal protein in corn event MON95275 is expressed by a Tripsacum dactyloides RCc3 promoter (United States Patent No. 9,617,553, SEQ ID NO: 13) and leader, operably linked to an enhancer derived from a Dalia mosaic virus promoter, Genbank accession EF513491, nucleotides 1 through 322; and a Setaria italica 14-3-3C protein gene intron (United States Patent Application Publication No. 2013-0031672 A2, SEQ ID NO: 151).

[0080] The Vip4Da2 protein in corn event MON95275 is expressed by a Zea mays Lipid Transfer Protein promoter and leader, enhanced with rearranged enhancer derived from multiple public Dahlia mosaic virus promoters and a Setaria italica Actin 4 gene intron (United States Patent Application No. 2013-0031672 A2, SEQ ID NO: 627). The Dahlia mosaic virus (DaMV) enhancer operably linked to the Zea mays Lipid Transfer Protein promoter and leader is a re-arranged composite of fragments derived from several public DaMV Genbank accessions, and is presented as SEQ ID NO: 24. A first fragment is derived from the promoter of the DaMV-Holland (DaMV- H) strain, Genbank accession EU090957, nucleotides 1177-1494. This fragment is operably linked to a second fragment derived from the DaMV-H promoter, nucleotides 1003-1176. Within the first fragment, relative to SEQ ID NO: 24, nucleotides 287 through 288, and nucleotides 319 through 322 were changed to sequences in analogous locations of a DaMV promoter within Genbank accession JX272320. In the native DaMV promoter configuration, the second fragmentwould precede the first fragment. The re-arrangement of these two fragments resulted in higher expression relative to the native fragment and was therefore selected for use in event MON95275.

[0081] The sequence encoding DvSnf7 specific dsRNA in com event MON95275 is driven by a promoter and leader derived from Cauliflower mosaic virus (CaMV) isolate NY8153 (presented as SEQ ID NO: 25), which is enhanced by an enhancer derived from the promoter of the pIIG gene encoding the physical impedance induced protein from Zea mays,' and a Zea mays hsp70 intron. The CaMV promoter / leader is derived from Genbank accession M90541, nucleotides 6,907 through 7,482. Relative to SEQ ID NO: 25, the second nucleotide was changed from a threonine (T) to an adenine (A) to remove a potential start codon in the operably linked cassette configuration. This CaMV promoter / leader comprised a longer leader sequence relative to the CaMV promoter and leader in corn event MON87411. This longer leader increased the expression levels of the DvSNF7 dsRNA in MON95275 relative to MON87411.

[0082] The expression of the Cry75Aal and Vip4Da2 transgene cassettes in MON95275 are oriented in a convergent manner as demonstrated in Fig. 1. The DvSnf7 transgene cassette in MON95275 is oriented in the divergent direction relative to the Cry75Aal transgene cassette, as demonstrated in Fig. 1. The DvSnf7 and the Cry75Aal transgene cassettes are separated from each other by an Intergenic Sequence Region (ISR4, United States Provisional Application Serial No. 62 / 875,752). Fig. 1 shows the relative positions of each element - enhancer (E), promoter (P), 5' UTR or leader (L), intron (I), 3' UTR (T), ISR4 (ISR), DvSnf7, Cry75Aal, and Vip4Da2 - comprised within SEQ ID NO: 9 and SEQ ID NO: 10.

[0083] As described herein, numerous constructs which varied in the use of expression elements, toxin coding sequences and orientation were evaluated. The construct used to create corn event MON95275 shown in Fig. 2 and presented as SEQ ID NO: 13, provided superior performance relative to other constructs when evaluated for resistance to Coleopteran insect pest infestation. In addition, corn event MON95275 is free of the markers used for selection of the transformed plant cell as a result of excision using Cre-recombinase. The CP4 selection cassette is shown in Fig. 2 and comprised within SEQ ID NO: 13. The CP4 selection cassette is flanked by two LoxP sites. Excision using Cre-recombinase resulted in the loss of the CP4 selection cassette after breeding with a Cre expressing transgenic corn event. The resulting progeny were evaluated for the absence of the selection cassette as well as the absence of the Cre-recombinase expression cassette, andthose progeny lacking both were selected for further evaluation, resulting in selection of the marker-free com event MON95275.

[0084] The event MON95275 was selected based on comparisons to thousands of different independent transgenic events, each transformed with a construct comprising the transgene cassette presented as SEQ ID NO: 13, or other constructs comprising the same or different toxins. The events generated expressing the insect toxins were compared to non-transgenic corn control plants of the same variety. The results as illustrated in the Examples show that the event MON95275 displayed superior properties due to expression of the Cry75Aal and Vip4Da2 protein, and the DvSnf7 specific dsRNA. The plurality of transgenic events produced using the construct used for generating the event MON95275 were each more likely than other events produced with other constructs to exhibit efficacious control of Coleopteran insect pests.

[0085] MON95275 was created through plant transformation techniques used to insert heterologous DNA (also known as transgenic DNA) randomly into a chromosome of the genome of a corn cell to produce a genetically engineered corn cell, also referred to as a “transgenic” or “recombinant” corn cell. Using this technique, many individual cells are transformed, each resulting in a unique “transgenic event” or “event” due to the random insertion of the foreign DNA into the genome. A transgenic plant is then regenerated from each individual transgenic cell. This results in every cell of the transgenic plant containing the uniquely inserted transgenic event as a stable part of its genome. This transgenic plant can then be used to produce seed which are then planted and grown into progeny plants, each containing the unique transgenic event.

[0086] Corn event MON95275 was produced by an Agrobacterium-mediated transformation process of com immature embryos with a single T-DNA binary system. In this system, an Agrobacterium strain employing one binary plasmid vector with a single T-DNA was utilized. The T-DNA construct comprised three transgene cassettes for the expression of the insect toxin coding sequences encoding Cry75Aa, Vip4Da2 and the dsRNA encoding sequence encoding DvSnf7, and a transgene cassette used for the selection of transformed com cells using glyphosate selection (CP4). The T-DNA constmetis SEQ ID NO: 13 and illustrated in Fig. 2 (“T-DNA Before Integration”). During integration, a single nucleotide was changed from a guanine (G) to a threonine (T) at nucleotide (nt) position 5,300 of SEQ ID NO: 13 (nt 4,986 of SEQ ID NO: 9 and nt 6,059 of SEQ ID NO: 10) in a region that is not within any of the coding sequences or expression elements. Also, during integration, six (6) nucleotides were inserted between the inserted T-DNAand 3' genomic flanking DNA and seven hundred forty-six (746) nucleotides were deleted from the wild-type genomic DNA. The glyphosate selection cassette was flanked on both sides with LoxP recognition sites which are recognized by Cre-recombinase, derived from Enterobacteria phage Pl (Larry Gilbertson (2003) Cre-lox recombination: Cre-active tools for plant biotechnology. TRENDS in Biotechnology, 21:12, 550-555)

[0087] As specifically described herein, corn event MON95275 was produced by a complex research and development process in which: (1) over one hundred sixty (160) plasmid vector constructs - which varied with respect to the coding sequences for the insecticidal proteins, the coding sequences for the transcriptional regulatory elements, and number and orientation of the cassettes within the constructs - were developed and transformed into com cells to create thousands of events that were tested and analyzed, resulting in the selection of the construct used to generate event MON95275; (2) thousands of com cells were transformed with the construct used to generate event MON95275, creating a population of transgenic plants in which each plant contained a unique transgenic event that was regenerated and tested; (3) the final event MON95275 was selected after a rigorous multi-year event selection process involving the testing and analysis of molecular characteristics, efficacy, protein expression, and agronomic properties in a variety of genetic backgrounds; and (4) the glyphosate selection cassette in corn event MON95275 was removed through in vivo Cre-excision to create a “marker-free” final event MON95275. Corn event MON95275 was thus produced and selected as a uniquely superior event useful for broadscale agronomic purposes.

[0088] The plasmid DNA inserted into the genome of corn event MON95275 was characterized by detailed molecular analysis. This analysis included: the insert number (number of integration sites within the com genome), the genomic insert location (the specific site in the corn genome where the insertion occurred), the copy number (the number of copies of the T-DNA within one locus), and the integrity of the transgenic inserted DNA. The detailed molecular analysis demonstrated that the integrated T-DNA containing the Cry75Aal, Vip4Da2, and DvSnf7 expression cassettes remained intact after integration and Cre-excision of the glyphosate (CP4) selection cassette. As used herein, an “expression cassette” or “cassette” is a recombinant DNA molecule comprising a combination of distinct elements that are to be expressed by a transformed cell. Table 1 provides a list of the elements contained in SEQ ID NO: 10, the DNA sequence that corresponds to com event MON95275.Table 1. Description of corn event MON95275

[0089] Corn event MON95275 is characterized as an insertion into a single locus in the corn genome, resulting in two new loci or junction sequences (e.g., sequences set forth in SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO :3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, and SEQ ID NO: 8) between the inserted DNA and the com genome DNA that are not known to appear naturally in the com genome or other transgenic corn events - they are unique to event MON95275. These junction sequences are useful in detecting the presence of the event MON95275 in corn cells, com tissue, com seed, and com plants or corn plant products, such as corn commodity products. DNA molecular probes and primer pairs are described herein that have been developed for use in identifying the presence of these various junction segments in biologicalsamples containing or suspected of containing com cells, com seed, com plant parts, or com plant tissue that contain the event MON95275.

[0090] As used herein, a “corn event MON95275 locus” refers to the genomic locus of the corn event MON95275 or a modified corn event MON95275 or a further modified corn event MON95275, wherein the com event MON95275 locus includes the flanking, junction and insertion sequences of the corn event MON95275 or the modified corn event MON95275 or further modified corn event MON95275. A modified corn event MON95275 or a further modified corn event MON95275 comprises one or more mutations, edits and / or genetic modifications in the corn event MON95275 locus, such as one or more mutations, edits and / or genetic modifications in a flanking, junction and / or insertion sequence(s) of the corn event MON95275 locus, relative to the com event MON95275.

[0091] According to present embodiments, a modified corn event MON95275 and methods of making a modified corn event MON95275 are provided. As is described further herein, various mutagenesis or targeted genome editing techniques and related tools are known and could be made or engineered to permit genetic modification or mutation of the transgenic insert, junction and / or the flanking genomic DNA of corn event MON95275, such as by deletion, insertion, transposition, inversion, and / or substitution of nucleic acid sequence(s), and / or by insertion or introduction of a guide RNA target site or a cognate target site or CgRRS, and the transgenic event as modified may still be uniquely characterized by the presence of heterologous DNA and / or one or more sequences of the insertion, junction(s) and / or flanking sequence(s) described herein at the same position or location in the genome previously occupied by the unmodified corn event MON95275 relative to flanking portions or sequences of the native corn genome. According to present embodiments, a modified transgenic event derived from corn event MON95275 may comprise all or part of the insertion sequence and / or transgene cassette of corn event MON95275, one or more of the junction sequence(s) of corn event MON95275, and / or one or more flanking sequence(s) of corn event MON95275 as described herein.

[0092] As used herein, a “modified corn event MON95275” refers to any genomic DNA or sequence of the com event MON95275 locus comprising one or more mutations, edits or genetic modifications relative to the genomic DNA or sequence of the corn event MON95275, wherein such mutations or edits are introduced or made by a mutagenesis or targeted genome editing technique of a com plant, plant part, tissue or cell comprising the corn event MON95275. A“modified corn event MON95275” includes, as a type of modified corn event MON95275, a “further modified com event MON95275” made by first inserting a target site or cognate target site or CgRRS into the corn event MON95275 locus and then further modifying the corn event MON95275 locus as described herein. For clarity, a “modified com event MON95275” includes genomic DNA or sequences of the corn event MON95275 locus comprising one or more mutations, edits or genetic modifications relative to the genomic DNA or sequence of the corn event MON95275, wherein such mutations or edits are introduced or made by a mutagenesis or targeted genome editing technique of a com plant, plant part, tissue or cell comprising the corn event MON95275, wherein such modified corn event MON95275 is not a further modified corn event MON95275. Methods and techniques of mutagenesis are known in the art and include, for example, chemical mutagenesis (z.e., treatment with a chemical mutagen, such as an azide, hydroxylamine, nitrous acid, acridine, nucleotide base analog, or alkylating agent - e.g., EMS (ethylmethane sulfonate), MNU (N-methyl-N-nitrosourea), etc.), physical mutagenesis (e.g., gamma rays, X-rays, UV, ion beam, other forms of radiation, etc.), and insertional mutagenesis (e.g., transposon or T-DNA insertion). As used herein, a “modified com plant” refers to a com or maize plant comprising a modified corn event MON95275 or a further modified corn event MON95275. Thus, a modified corn plant part, plant seed, plant tissue, or plant cell comprising a modified corn event MON95275 or a further modified corn event MON95275 that is derived, taken or descended from a modified com plant and / or created by genetic modification, mutation or editing of the transgenic insert, junction and / or the flanking genomic DNA of com event MON95275 in a com plant part, plant seed, plant tissue, or plant cell using a mutagenesis or targeted genome editing technique.

[0093] As used herein, a “target site” for genome editing refers to the location of a polynucleotide sequence within a plant genome, which may be within or near corn event MON95275, that is bound and cleaved by a site-specific nuclease introducing a double stranded break (or singlestranded nick) into the nucleic acid backbone of the polynucleotide sequence and / or its complementary DNA strand. A target site may comprise at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 26, at least 27, at least 29, or at least 30 consecutive nucleotides. A “target site” for a RNA-guided nuclease may comprise the sequence of either complementary strand of a double-stranded nucleic acid (DNA) molecule or chromosomeat the target site that is bound or hybridized to a guide RNA of a ribonucleoprotein complex comprising the RNA-guided nuclease. A site-specific nuclease may bind to a target site, such as via a non-coding guide RNA (e.g., without being limiting, a CRISPR RNA (crRNA) or a singleguide RNA (sgRNA) as described further below). A non-coding guide RNA (gRNA) provided herein may be complementary to a target site (e.g., complementary to either strand of a doublestranded nucleic acid molecule or chromosome at the target site). It will be appreciated that perfect identity or complementarity may not be required for a non-coding guide RNA to bind or hybridize to a target site. For example, at least 1, at least 2, at least 3, at least 4, at least 5, at least 6, at least 7, or at least 8 mismatches (or more) between a target site and a non-coding RNA may be tolerated. A “target site” also refers to the location of a polynucleotide sequence within a plant genome, which may be within or near corn event MON95275 or a modified corn event MON95275, that is bound and cleaved by another site-specific nuclease that may not be guided by a non-coding RNA molecule, such as a meganuclease, zinc finger nuclease (ZFN), or a transcription activator-like effector nuclease (TALEN), to introduce a double stranded break (or single-stranded nick) into the polynucleotide sequence and / or its complementary DNA strand. As used herein, a “target region” or a “targeted region” refers to a polynucleotide sequence or region that is flanked by two or more target sites. Without being limiting, in some embodiments a target region may be subjected to a mutation, deletion, insertion or inversion. As used herein, “flanked” when used to describe a target region of a polynucleotide sequence or molecule, refers to two or more target sites of the polynucleotide sequence or molecule surrounding the target region, with one target site on each side of the target region.

[0094] As used herein, a “donor molecule”, “donor template”, or “donor template molecule” (collectively a “donor template”), which may be a recombinant DNA donor template, is defined as a nucleic acid molecule having a nucleic acid template or insertion sequence for site-directed, targeted insertion or recombination into the genome of a plant cell via repair of a nick or doublestranded DNA break in the genome of a plant cell. For example, a “donor template” may be used for site-directed integration of a guide RNA target site or a cognate target site or CgRRS into a target site within the genome of a plant. A targeted genome editing technique provided herein may comprise the use of one or more, two or more, three or more, four or more, or five or more donor molecules or templates. A “donor template” may be a single-stranded or double-stranded DNA or RNA molecule or plasmid. An “insertion sequence” of a donor template is a sequence designedfor targeted insertion into the genome of a plant cell, which may be of any suitable length, such as to include a guide RNA target site or a cognate target site or CgRRS. Such an insertion sequence of a donor template is distinct and different from the transgenic insertion or insert of corn event MON95275, although they may potentially have sequence(s) in common. A donor template may also have at least one homology sequence or homology arm, such as two homology arms, to direct the integration of a mutation or insertion sequence into a target site within the genome of a plant via homologous recombination, wherein the homology sequence or homology arm(s) are identical or complementary, or have a percent identity or percent complementarity, to a sequence at or near the target site within the genome of the plant. When a donor template comprises homology arm(s) and an insertion sequence, the homology arm(s) will flank or surround the insertion sequence of the donor template.

[0095] As used herein, a “targeted genome editing technique” refers to any method, protocol, or technique that can be used to make a targeted mutation or edit, such as one or more insertions, deletions, substitutions, inversions, transpositions, mutations and / or other genetic modifications at or near a target site in the genome of a plant, and / or a deletion or excision of a target region between two target sites in the genome of a plant, using a site-specific nuclease, such as a meganuclease, a zinc-finger nuclease (ZFN), an RNA-guided endonuclease (e.g., the CRISPR / Cas9 system), a TALE-endonuclease (TALEN), a recombinase, or a transposase. A site-specific nuclease may introduce a double stranded break (or single-stranded nick) into the nucleic acid backbone of the polynucleotide sequence and / or its complementary DNA strand. Following the introduction of the single or double-stranded break or nick or cleavage of DNA at or near the target site(s) by the sitespecific nuclease, the genomic sequence can be repaired via a double strand break repair pathway, which may include, for example, non-homologous end-joining (NHEJ), microhomology-mediated end joining (MMEJ), homologous recombination, synthesis-dependent strand annealing (SDSA), single-strand annealing (SSA), or a combination of any thereof, at or near the target site(s). However, if the repair is imperfect, a mutation or edit may be introduced at or near the target site(s), and a target region between two or more target sites may be deleted or excised.

[0096] A “site-specific nuclease” provided herein may be selected from the group consisting of a zinc-finger nuclease (ZFN), a meganuclease, an RNA-guided endonuclease, a TALE- endonuclease (TALEN), a recombinase, a transposase, or any combination thereof. See, e.g., Khandagale, K. et al., “Genome editing for targeted improvement in plants,” Plant Biotechnol Rep10: 327-343 (2016); and Gaj, T. et al., “ZFN, TALEN and CRISPR / Cas-based methods for genome engineering,” Trends BiotechnoL 31(7): 397-405 (2013), the contents and disclosures of which are incorporated herein by reference. A recombinase may be a serine recombinase attached to a DNA recognition motif, a tyrosine recombinase attached to a DNA recognition motif or other recombinase enzyme known in the art. A recombinase or transposase may be a DNA transposase or recombinase attached to a DNA binding domain. A tyrosine recombinase attached to a DNA recognition motif may be selected from the group consisting of a Cre recombinase, a Flp recombinase, and a Tnpl recombinase. According to some embodiments, a Cre recombinase or a Gin recombinase provided herein is tethered to a zinc-finger DNA binding domain. In another embodiment, a serine recombinase attached to a DNA recognition motif provided herein is selected from the group consisting of a PhiC31 integrase, an R4 integrase, and a TP-901 integrase. In another embodiment, a DNA transposase attached to a DNA binding domain provided herein is selected from the group consisting of a TALE-piggyBac and TALE-Mutator.

[0097] According to embodiments of the present disclosure, an RNA-guided endonuclease may be selected from the group consisting of Casl, CaslB, Cas2, Cas3, Cas4, Cas5, Cas6, Cas7, Cas8, Cas9 (also known as Csnl and Csxl2), CaslO, Csyl, Csy2, Csy3, Csel, Cse2, Cscl, Csc2, Csa5, Csn2, Csm2, Csm3, Csm4, Csm5, Csm6, Cmrl, Cmr3, Cmr4, Cmr5, Cmr6, Csbl, Csb2, Csb3, Csxl7, Csxl4, CsxlO, Csxl6, CsaX, Csx3, Csxl, Csxl5, Csfl, Csf2, Csf3, Csf4, Casl2a, Cpfl, CasX, CasY, and homologs or modified versions thereof, Argonaute (non-limiting examples of Argonaute proteins include Thermus thermophilus Argonaute (TtAgo), Pyrococcus furiosus Argonaute (PfAgo), Natronobacterium gregoryi Argonaute (NgAgo) and homologs or modified versions thereof. According to some embodiments, an RNA-guided endonuclease may be a Cas9 or Casl 2a or Cpfl enzyme.

[0098] In an aspect, a site-specific nuclease provided herein is selected from the group consisting of a zinc-finger nuclease, a meganuclease, an RNA-guided nuclease, a TALE-nuclease, a recombinase, a transposase, or any combination thereof. In another aspect, a site-specific nuclease provided herein is selected from the group consisting of a Cas9 or a Casl2a or Cpfl. In another aspect, a site-specific nuclease provided herein is selected from the group consisting of a Casl, a CaslB, a Cas2, a Cas3, a Cas4, a Cas5, a Cas6, a Cas7, a Cas8, a Cas9, a CaslO, a Casl2a, a Csyl, a Csy2, a Csy3, a Csel, a Cse2, a Cscl, a Csc2, a Csa5, a Csn2, a Csm2, a Csm3, a Csm4, a Csm5, a Csm6, a Cmrl, a Cmr3, a Cmr4, a Cmr5, a Cmr6, a Csbl, a Csb2, a Csb3, a Csxl7, a Csxl4, aCsxlO, a Csxl 6, a CsaX, a Csx3, a Csxl, a Csxl 5, a Csfl , a Csf2, a Csfl, a Csf4, a Cpfl, CasX, CasY, a homolog thereof, or a modified version thereof. In another aspect, an RNA-guided nuclease provided herein is selected from the group consisting of a Cas9 or a Casl2a or Cpfl. In another aspect, an RNA guided nuclease provided herein is selected from the group consisting of a Casl, a CaslB, a Cas2, a Cas3, a Cas4, a Cas5, a Cas6, a Cas7, a Cas8, a Cas9, a CaslO, a Casl2a, a Csyl, a Csy2, a Csy3, a Csel, a Cse2, a Cscl, a Csc2, a Csa5, a Csn2, a Csm2, a Csm3, a Csm4, a Csm5, a Csm6, a Cmrl, a Cmr3, a Cmr4, a Cmr5, a Cmr6, a Csbl, a Csb2, a Csb3, a Csxl7, a Csxl4, a CsxlO, a Csxl6, a CsaX, a Csx3, a Csxl, a Csxl5, a Csfl, a Csf2, a Csf3, a Csf4, a Cpfl, CasX, CasY, a homolog thereof, or a modified version thereof. In another aspect, a method and / or a composition provided herein comprises at least one, at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten sitespecific nucleases. In yet another aspect, a method and / or a composition provided herein comprises at least one, at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten polynucleotides encoding at least one, at least two, at least three, at least four, at least five, at least six, at least seven, at least eight, at least nine, or at least ten site-specific nucleases.

[0099] For RNA-guided endonucleases, a guide RNA (gRNA) molecule is further provided to direct the endonuclease to a target site in the genome of the plant via base-pairing or hybridization to cause a DSB or nick at or near the target site. The gRNA may be transformed or introduced into a plant cell or tissue (perhaps along with a nuclease, or nuclease-encoding DNA molecule, construct or vector) as a gRNA molecule, or as a recombinant DNA molecule, construct, or vector comprising a polynucleotide or transcribable DNA sequence encoding the guide RNA operably linked to a plant-expressible promoter. As understood in the art, a “guide RNA” may comprise, for example, a CRISPR RNA (crRNA), a single-chain guide RNA (sgRNA), or any other RNA molecule that may guide or direct an endonuclease to a specific target site in the genome. A “single-chain guide RNA” (or “sgRNA”) is a RNA molecule comprising a crRNA covalently linked a tracrRNA by a linker sequence, which may be expressed as a single RNA transcript or molecule. The guide RNA comprises a guide or targeting sequence that is identical or complementary to a target site within the plant genome, such as within or near corn event MON95275 or a modified corn event MON95275. A protospacer-adjacent motif (PAM) may be present in the genome immediately adjacent and upstream or downstream of the genomic targetsite sequence complementary to the targeting sequence of the guide RNA as known in the art. See, e.g., Wu, X. et al., “Target specificity of the CRISPR-Cas9 system,” Quant Biol. 2(2): 59-70 (2014), the content and disclosure of which is incorporated herein by reference. The guide RNA may typically be a non-coding RNA molecule that does not encode a protein. The guide sequence of the guide RNA may be at least 10 nucleotides in length, such as 12-40 nucleotides, 12-30 nucleotides, 12-20 nucleotides, 12-35 nucleotides, 12-30 nucleotides, 15-30 nucleotides, 17-30 nucleotides, or 17-25 nucleotides in length, or about 12, 13, 14, 15, 16, 17, 18, 19, 20, 21, 22, 23, 24, 25 or more nucleotides in length. The guide sequence may be at least 95%, at least 96%, at least 97%, at least 99% or 100% identical or complementary to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, or more consecutive nucleotides of a DNA sequence at the genomic target site. As used herein, the term “consecutive” in reference to a polynucleotide or protein sequence means without deletions or gaps in the sequence.

[0100] In addition to the guide sequence, a guide RNA may further comprise one or more other structural or scaffold sequence(s), which may bind or interact with an RNA-guided endonuclease. Such scaffold or structural sequences may further interact with other RNA molecules (e.g., tracrRNA). Methods and techniques for designing targeting constructs and guide RNAs for genome editing and site-directed integration at a target site within the genome of a plant using an RNA-guided endonuclease are known in the art.

[0101] According to some embodiments, recombinant DNA molecules, constructs and vectors are provided comprising a polynucleotide or transcribable DNA sequence encoding a site-specific nuclease, such as a zinc-finger nuclease (ZFN), a meganuclease, an RNA-guided endonuclease, a TALE-endonuclease (TALEN), a recombinase, or a transposase, wherein the coding sequence is operably linked to a plant expressible promoter. For RNA-guided endonucleases, recombinant DNA molecules, constructs and vectors are further provided comprising a polynucleotide or transcribable DNA sequence encoding a guide RNA, wherein the guide RNA comprises a guide sequence of sufficient length having a percent identity or complementarity to a target site within the genome of a plant. According to some embodiments, recombinant DNA molecules, constructs and vectors are provided comprising a first polynucleotide or transcribable DNA sequence encoding a site-specific nuclease and a second polynucleotide or transcribable DNA sequence encoding one or more gRNAs. According to some embodiments, each polynucleotide ortranscribable DNA sequence of a recombinant DNA molecule, construct and vector that encodes a site-specific nuclease and / or a guide RNA may be operably linked to a plant expressible promoter, such as an inducible promoter, a constitutive promoter, a tissue-specific promoter, etc.

[0102] According to some embodiments, recombinant DNA molecules, constructs and vectors are provided comprising a polynucleotide or transcribable DNA sequence encoding a site-specific nuclease, such as a zine-finger nuclease (ZFN), a meganuclease, an RNA-guided endonuclease, a TALE-endonuclease (TALEN), a recombinase, or a transposase, wherein the coding sequence is operably linked to a plant expressible promoter. For RNA-guided endonucleases, recombinant DNA molecules, constructs and vectors are further provided comprising a polynucleotide or transcribable DNA sequence encoding a guide RNA, wherein the guide RNA comprises a guide sequence of sufficient length having a percent identity or complementarity to a target site within the genome of a plant. According to some embodiments, recombinant DNA molecules, constructs and vectors are provided comprising a first polynucleotide or transcribable DNA sequence encoding a site-specific nuclease and a second polynucleotide or transcribable DNA sequence encoding one or more gRNAs. According to some embodiments, each polynucleotide or transcribable DNA sequence of a recombinant DNA molecule, construct and vector that encodes a site-specific nuclease and / or a guide RNA may be operably linked to a plant expressible promoter, such as an inducible promoter, a constitutive promoter, a tissue-specific promoter, etc.

[0103] According to some embodiments, a recombinant DNA molecule, construct or vector may comprise a first polynucleotide sequence encoding a site-specific nuclease and a second polynucleotide sequence encoding a guide RNA(s) that may be introduced into a plant cell together via plant transformation techniques. Alternatively, two recombinant DNA molecules, constructs or vectors may be provided including a first recombinant DNA molecule, construct or vector and a second DNA molecule, construct or vector that may be introduced into a plant cell together or sequentially via plant transformation techniques, wherein the first recombinant DNA molecule, construct or vector comprises a polynucleotide sequence encoding a site-specific nuclease and the second recombinant DNA molecule, construct or vector comprises a polynucleotide sequence encoding a guide RNA(s). According to some embodiments, a recombinant DNA molecule, construct or vector comprising a polynucleotide sequence encoding a site-specific nuclease may be introduced via plant transformation techniques into a plant cell that already comprises (or is transformed with) a recombinant DNA construct or vector comprising a polynucleotide sequenceencoding a guide RNA(s). Alternatively, a recombinant DNA molecule, construct or vector comprising a polynucleotide sequence encoding a guide RNA may be introduced via plant transformation techniques into a plant cell that already comprises (or is transformed with) a recombinant DNA construct or vector comprising a polynucleotide sequence encoding a sitespecific nuclease. According to yet further embodiments, a first plant comprising (or transformed with) a recombinant DNA construct or vector comprising a polynucleotide sequence encoding a site-specific nuclease may be crossed with a second plant comprising (or transformed with) a recombinant DNA construct or vector comprising a polynucleotide sequence encoding a guide RNA(s). Such recombinant DNA molecules, constructs or vectors may be transiently transformed into a plant cell or stably transformed or more preferably integrated into the genome of a plant cell.

[0104] In an aspect, molecules or vectors comprising polynucleotides encoding a site-specific nuclease, and optionally one or more, two or more, three or more, or four or more gRNAs are provided to a plant cell by transformation methods known in the art (e.g., without being limiting, particle bombardment, PEG-mediated protoplast transfection or Agrobacterium-mediated transformation). In an aspect, molecules or vectors comprising polynucleotides encoding a Cas9 nuclease, and optionally one or more, two or more, three or more, or four or more gRNAs are provided to a plant cell by transformation methods known in the art e.g., without being limiting, particle bombardment, PEG-mediated protoplast transfection or Agrobacterium-mediated transformation). In another aspect, vectors comprising polynucleotides encoding a Cpfl and, optionally one or more, two or more, three or more, or four or more crRNAs are provided to a cell by transformation methods known in the art (e.g., without being limiting, viral transfection, particle bombardment, PEG-mediated protoplast transfection or Agrobacterium-mediated transformation).

[0105] Several site-specific nucleases, such as recombinases, zinc finger nucleases (ZFNs), meganucleases, and TALENs, are not RNA-guided and instead rely on their protein structure to determine their target site for causing the DSB or nick, or they are fused, tethered, or attached to a DNA-binding protein domain or motif. The protein structure of the site-specific nuclease (or the fused / attached / tethered DNA binding domain) may target the site-specific nuclease to the target site. According to many of these embodiments, non-RNA-guided site-specific nucleases, such as recombinases, zinc finger nucleases (ZFNs), meganucleases, and TALENs, may be designed, engineered and constructed according to known methods to target and bind to a target site in thegenome of a plant, to create a DSB or nick at or near such genomic target site or locus. For example, an engineered site-specific nuclease, such as a recombinase, zinc finger nuclease (ZFN), meganuclease, or TALEN, may be designed to target and bind to a genomic target site within the genome of a plant to create a DSB or nick at the genomic target site.

[0106] In an aspect, a targeted genome editing technique described herein may comprise the use of a zinc finger nuclease (ZFN). ZFNs are synthetic proteins consisting of an engineered zinc finger DNA-binding domain fused to a cleavage domain (or a cleavage half-domain), which may be derived from a restriction endonuclease (e.g., FokI). The DNA binding domain may be canonical (C2H2) or non-canonical (e.g., C3H or C4). The DNA-binding domain can comprise one or more zinc fingers (e.g., 2, 3, 4, 5, 6, 7, 8, 9 or more zinc fingers) depending on the target site. Multiple zinc fingers in a DNA-binding domain may be separated by linker sequence(s). ZFNs can be designed to cleave almost any stretch of double-stranded DNA by modification of the zinc finger DNA-binding domain. ZFNs form dimers from monomers composed of a nonspecific DNA cleavage domain (e.g, derived from the FokI nuclease) fused to a DNA-binding domain comprising a zinc finger array engineered to bind a target site DNA sequence. The DNA- binding domain of a ZFN may typically be composed of 3-4 (or more) zinc-fingers. The amino acids at positions -1, +2, +3, and +6 relative to the start of the zinc finger a-helix, which contribute to site-specific binding to the target site, can be changed and customized to fit specific target sequences. The other amino acids may form a consensus backbone to generate ZFNs with different sequence specificities. Methods and rules for designing ZFNs for targeting and binding to specific target sequences are known in the art. See, e.g., US Patent App. Nos. 2005 / 0064474, 2009 / 0117617, and 2012 / 0142062, the contents and disclosures of which are incorporated herein by reference. The FokI nuclease domain may require dimerization to cleave DNA and therefore two ZFNs with their C-terminal regions are needed to bind opposite DNA strands of the cleavage site (separated by 5-7 bp). The ZFN monomer can cut the target site if the two-ZF-binding sites are palindromic. A ZFN, as used herein, is broad and includes a monomeric ZFN that can cleave double stranded DNA without assistance from another ZFN. The term ZFN may also be used to refer to one or both members of a pair of ZFNs that are engineered to work together to cleave DNA at the same site.

[0107] Without being limited by any scientific theory, because the DNA-binding specificities of zinc finger domains can be re-engineered using one of various methods, customized ZFNs cantheoretically be constructed to target nearly any target sequence (e.g., at or near com event MON95275 in a plant genome). Publicly available methods for engineering zinc finger domains include Context-dependent Assembly (CoDA), Oligomerized Pool Engineering (OPEN), and Modular Assembly. In an aspect, a method and / or composition provided herein comprises one or more, two or more, three or more, four or more, or five or more ZFNs. In another aspect, a ZFN provided herein is capable of generating a targeted DSB or nick. In an aspect, vectors comprising polynucleotides encoding one or more, two or more, three or more, four or more, or five or more ZFNs are provided to a cell by transformation methods known in the art (e.g., without being limiting, viral transfection, particle bombardment, PEG-mediated protoplast transfection, or Agrobacterium-mediated transformation). The ZFNs may be introduced as ZFN proteins, as polynucleotides encoding ZFN proteins, and / or as combinations of proteins and protein-encoding polynucleotides.

[0108] In an aspect, a targeted genome editing technique described herein may comprise the use of a meganuclease. Meganucleases, which are commonly identified in microbes, such as the LAGLID ADG family of homing endonucleases, are unique enzymes with high activity and long recognition sequences (> 14 bp) resulting in site-specific digestion of target DNA. Engineered versions of naturally occurring meganucleases typically have extended DNA recognition sequences (for example, 14 to 40 bp). According to some embodiments, a meganuclease may comprise a scaffold or base enzyme selected from the group consisting of I-Crel, I-Ceul, I-Msol, I-Scel, I-Anil, and I-Dmol. The engineering of meganucleases can be more challenging than ZFNs and TALENs because the DNA recognition and cleavage functions of meganucleases are intertwined in a single domain. Specialized methods of mutagenesis and high-throughput screening have been used to create novel meganuclease variants that recognize unique sequences and possess improved nuclease activity. Thus, a meganuclease may be selected or engineered to bind to a genomic target site or sequence in a plant. In an aspect, a method and / or composition provided herein comprises one or more, two or more, three or more, four or more, or five or more meganucleases. In another aspect, a meganuclease provided herein is capable of generating a targeted DSB. In an aspect, vectors comprising polynucleotides encoding one or more, two or more, three or more, four or more, or five or more meganucleases are provided to a cell by transformation methods known in the art (e.g., without being limiting, viral transfection, particle bombardment, PEG-mediated protoplast transfection or Agrobacterium-mediated transformation).

[0109] In an aspect, a targeted genome editing technique described herein may comprise the use of a transcription activator-like effector nuclease (TALEN). TALENs are artificial restriction enzymes generated by fusing the transcription activator-like effector (TALE) DNA binding domain to a nuclease domain (e.g., FokT). In some aspects, the nuclease is selected from a group consisting of PvuII, MutH, TevI, FokI, AhvI, Mlyl, Sbfl, Sdal, Sisi, CleDORF, Clo051, and PeptO7I. For FokI nuclease, when each member of a TALEN pair binds to the DNA sites flanking a target site, the FokI monomers dimerize and cause a double-stranded DNA break at the target site. Besides the wild-type FokI cleavage domain, variants of the FokI cleavage domain with mutations have been designed to improve cleavage specificity and cleavage activity. The FokI domain functions as a dimer, requiring two constructs with unique DNA binding domains for sites in the target genome with proper orientation and spacing. Both the number of amino acid residues between the TALEN DNA binding domain and the FokI cleavage domain and the number of bases between the two individual TALEN binding sites are parameters for achieving high levels of activity. The term TALEN, as used herein, is broad and includes a monomeric TALEN that can cleave double stranded DNA without assistance from another TALEN. The term TALEN also refers to one or both members of a pair of TALENs that work together to cleave DNA at the same site.

[0110] Besides the wild-type FokI cleavage domain, variants of the FokI cleavage domain with mutations have been designed to improve cleavage specificity and cleavage activity. The FokI domain functions as a dimer, requiring two constructs with unique DNA binding domains for sites in the target genome with proper orientation and spacing. Both the number of amino acid residues between the TALEN DNA binding domain and the FokI cleavage domain and the number of bases between the two individual TALEN binding sites are parameters for achieving high levels of activity. PvuII, MutH, and TevI cleavage domains are useful alternatives to FokI and FokI variants for use with TALEs. PvuII functions as a highly specific cleavage domain when coupled to a TALE (see Yank et al. 2013. PLoS One. 8: e82539). MutHis capable of introducing strand-specific nicks in DNA (see Gabsalilow et al. 2013. Nucleic Acids Research. 41: e83). TevI introduces doublestranded breaks in DNA at targeted sites (see Beurdeley et al., 2013. Nature Communications. 4: 1762).

[0111] Transcription activator-like effectors (TALEs) can be engineered to bind practically any DNA sequence, such as at or near the genomic locus of corn event MON95275 in a plant. TALEhas a central DNA-binding domain composed of 13-28 repeat monomers of 33-34 amino acids. The amino acids of each monomer are highly conserved, except for hypervariable amino acid residues at positions 12 and 13. The two variable amino acids are called repeat- variable diresidues (RVDs). The amino acid pairs NI, NG, HD, and NN of RVDs preferentially recognize adenine, thymine, cytosine, and guanine / adenine, respectively, and modulation of RVDs can recognize consecutive DNA bases. This simple relationship between amino acid sequence and DNA recognition has allowed for the engineering of specific DNA binding domains by selecting a combination of repeat segments containing the appropriate RVDs. The relationship between amino acid sequence and DNA recognition of the TALE binding domain allows for designable proteins. Software programs such as DNA Works can be used to design TALE constructs. Other methods of designing TALE constructs are known to those of skill in the art. See Doyle el al., Nucleic Acids Research (2012) 40: W117-122.; Cermak el al., Nucleic Acids Research (2011). 39:e82; and tale-nt.cac.cornell.edu / about. In an aspect, a method and / or composition provided herein comprises one or more, two or more, three or more, four or more, or five or more TALENs. In another aspect, a TALEN provided herein is capable of generating a targeted DSB. In an aspect, vectors comprising polynucleotides encoding one or more, two or more, three or more, four or more, or five or more TALENs are provided to a cell by transformation methods known in the art (e.g., without being limiting, viral transfection, particle bombardment, PEG-mediated protoplast transfection or Agrobacterium-mediated transformation). See, e.g., US Patent App. Nos. 2011 / 0145940, 2011 / 0301073, and 2013 / 0117869, the contents and disclosures of which are incorporated herein by reference.

[0112] In an aspect, a targeted genome editing technique described herein may comprise the use of a recombinase. In some embodiments, a tyrosine recombinase attached, etc., to a DNA recognition domain or motif may be selected from the group consisting of a Cre recombinase, a Flp recombinase, and a Tnp l recombinase. In an aspect, a Cre recombinase or a Gin recombinase provided herein may be tethered to a zinc-finger DNA binding domain. The Flp- / ’ ? / ' site-directed recombination system may come from the 2p plasmid from the baker’s yeast Saccharomyces cerevisiae. In this system, Flp recombinase (flippase) may recombine sequences between flippase recognition target (FRT) sites. FRT sites comprise 34 nucleotides. Flp may bind to the “arms” of the FRT sites (one arm is in reverse orientation) and cleaves the FRT site at either end of an intervening nucleic acid sequence. After cleavage, Flp may recombine nucleic acid sequencesbetween two FRT sites. Cre-lox is a site-directed recombination system derived from the bacteriophage Pl that is similar to the Flp- / ’7 7' recombination system. Cre-lox can be used to invert a nucleic acid sequence, delete a nucleic acid sequence, or translocate a nucleic acid sequence. In this system, Cre recombinase may recombine a pair of lox nucleic acid sequences. Lox sites comprise 34 nucleotides, with the first and last 13 nucleotides (arms) being palindromic. During recombination, Cre recombinase protein binds to two lox sites on different nucleic acids and cleaves at the lox sites. The cleaved nucleic acids are spliced together (reciprocally translocated) and recombination is complete. In another aspect, a lox site provided herein is a loxP, lox 2272, loxN, lox 511, lox 5171, lox71, lox66, M2, M3, M7, or Mi l site.

[0113] As used herein, the term “derived” or “derived from” in reference to a particular DNA molecule, amplicon or sequence in relation to a corn cell, corn tissue, corn seed, corn plant, corn plant part and / or com plant product, such as a com commodity product, means that the DNA molecule, amplicon or sequence is taken, purified, isolated, or made, directly or indirectly, from such com cell, corn tissue, com seed, corn plant, com plant part and / or corn plant product, such as a com commodity product, as the case may be. Alternatively, the term “derived” or “derived from” in reference to a corn plant product, such as a corn commodity product, in relation to a corn cell, corn tissue, corn seed, corn plant, and / or com plant part, means that the com plant product is taken, purified, isolated, or made, directly or indirectly, from such com cell, corn tissue, corn seed, corn plant, and / or corn plant part, as the case may be. “Capable of being detected” refers to the ability of a particular DNA molecule, segment, or sequence to be detected in a sample, such as by amplification and determining its presence, size, or sequence such as by DNA sequence analysis, and / or binding of a probe to the target DNA molecule, segment, or sequence.

[0114] A “sample” is intended to refer to any composition comprising or derived from, either directly or indirectly, a biological sample, source or material. The sample may generally comprise corn DNA and / or substantially or completely pure, purified or isolated com DNA. A “biological sample” contains biological materials, including but not limited to DNA obtained or derived from, either directly or indirectly, the genome of a com cell(s), corn tissue(s), com seed(s), corn plant(s), corn plant part(s) and / or corn plant product(s), such as a com commodity product(s). Such corn cell(s), com tissue(s), corn seed(s), corn plant(s), corn plant part(s) and / or corn plant product(s), such as a corn commodity product(s), may comprise com event MON95275 or a modified corn event MON95275 or DNA molecule(s) and / or DNA segment(s) comprising com eventMON95275 or a modified com event MON95275. Tn some embodiments, a sample or biological sample may comprise corn cell(s), corn tissue(s), corn seed(s), corn plant(s), corn plant part(s), and / or corn plant product(s), whose cells or cellular membranes have been fractured (e.g., disrupted or opened) to release the contents of the com cell(s) including genomic DNA and / or make the contents of the com cell(s) including genomic DNA accessible or usable for assays or testing. “Directly” refers to directly obtaining DNA by a skilled artisan from the corn genome by fracturing com cells (or by obtaining samples of com that contain fractured com cells) and exposing or using the genomic DNA from com cells for the purposes of detection. “Indirectly” refers to obtaining by a skilled artisan a target or specific reference DNA ( / . , a novel and unique junction segment(s) described herein as being diagnostic for the presence of the corn event MON95275 or a modified corn event MON95275) in a particular sample, by means other than by obtaining directly via fracturing of corn cells or obtaining a sample of corn that contains fractured corn cells. Such indirect means include, but are not limited to, amplification of a DNA segment that contains a DNA sequence targeted by a particular probe(s) and / or primer set(s) designed to bind with specificity to or near the target sequence, or amplification of a DNA segment comprising all or part of a target sequence that can be measured and characterized (e.g., measured by migration or separation from other segments of DNA and / or identification in an effective matrix, such as an agarose or acrylamide gel or the like, or characterized by direct sequence analysis of the amplicon(s), or cloning of the amplicon(s) into a vector(s) and direct sequencing of the inserted amplicon(s) present within such vector(s).

[0115] Detailed molecular analysis also demonstrated that event MON95275 contains a single T- DNA insertion with one copy of each of the Cry75Aal, Vip4Da2, and DvSnf7 specific dsRNA expression cassettes. No additional elements from the transformation construct other than portions of the Agrobacterium tumefaciens left and right border regions used for transgenic DNA transfer from the plant transformation plasmid to the corn genome were identified in event MON95275. Further, thermal amplification producing specific amplicons diagnostic for the presence of event MON95275 in a sample and DNA sequence analyses were performed to determine the arbitrarily assigned 5' and 3' insert-to-plant genome junctions, confirm the organization of the elements within the insert, and determine the complete DNA sequence of the inserted transgenic DNA (SEQ ID NO: 9). SEQ ID NO: 11 is a sequence representing the one thousand seventy-three (1,073) base-pair (bp) 5' LH244 corn genomic DNA sequence flanking the inserted T-DNA sequencepresented as SEQ ID NO: 9. SEQ ID NO: 12 is a sequence representing the one thousand one hundred six (1,106) bp 3' LH244 corn genomic DNA sequence flanking the inserted T-DNA sequence presented as SEQ ID NO: 9. SEQ ID NO: 7 is a sequence representing the one thousand two hundred twenty-six (1,226) base-pair (bp) 5' LH244 corn genomic DNA sequence flanking the inserted T-DNA sequence combined with one hundred fifty-three (153) bp of inserted T-DNA sequence presented as SEQ ID NO: 9. SEQ ID NO: 8 is a sequence representing one hundred one (lOl) bp of inserted T -DNA sequence with the one thousand one hundred six ( 1 , 106) bp 3 ' LH244 corn genomic DNA sequence flanking the inserted T-DNA sequence presented as SEQ ID NO: 9. SEQ ID NO: 10 corresponds to corn event MON95275 and contains a contiguous sequence (contig) comprising the 5' LH244 flanking sequence, the transgene insert of MON95275, and the 3 ' LH244 flanking sequence, and thus contains the insert-to-plant genome junction sequences.

[0116] Unless otherwise noted herein, terms are to be understood according to conventional usage by those of ordinary skill in the relevant art. Definitions of common terms in molecular biology may be found in Rieger etal., Glossary of Genetics: Classical and Molecular, 5thedition, Springer- Verlag: New York, 1991; and Lewin, Genes V, Oxford University Press: New York, 1994, along with other sources known to those of ordinary skill in the art. As used herein, the term “corn” means species belong to the genus Zea, preferably Zea mays and includes all plant varieties that can be bred with corn plants containing event MON95275, including wild corn species as well as those plants belonging to the genus Zea that permit breeding between species.

[0117] The present invention provides for transgenic plants which have been transformed with a DNA construct that contains expression cassettes expressing toxic amounts of the insecticidal proteins Cry75Aal, Vip4Da2, and the DvSnf7 specific dsRNA. What is meant by toxic amount is an efficacious amount, an insecticidal amount, an insecticidally-effective amount, a target insect suppressive amount, an efficacious pesticidal amount, an amount in the diet of insects in the order of Coleoptera that is insecticidal, and other similar terms to be understood according to conventional usage by those of ordinary skill in the relevant art. Corn plants transformed according to the methods and with the DNA construct disclosed herein are resistant to Coleopteran insect pests.

[0118] A transgenic plant is produced by transformation of a plant cell with a recombinant DNA construct that includes the expression cassette(s) and transgene(s) as described herein, which is also heterologous with respect to the plant cell, regeneration of a plant resulting from the insertionof the transgene into the genome of the plant cell, and selection of a particular plant characterized by insertion into a particular genome location and the number of efficacious features of the regenerated transgenic plant. The term “transgenic event” or “event” refers to the inserted transgenic DNA in the plant genome and flanking genomic sequences immediately adjacent to the inserted transgenic DNA in the genome of the transformed plant, but also refers to a DNA molecule comprising the inserted transgenic DNA in the plant genome and flanking genomic sequences. Each event is unique and would be expected to be transferred to progeny plants that receive the transgenic DNA and event through genetic inheritance and / or segregation from a parent as the result of a sexual or self-cross of a first parental line that includes the inserted transgenic DNA and event either with itself or a second parental line that may or may not contain the same transgenic DNA and event. The parental line that includes the inserted transgenic DNA and event may itself be the original transformant or a progeny plant of said original transformant that may have been generated by “selfing” the transformant with itself or crossing the transformant or a progeny plant of the transformant that includes the inserted transgenic DNA and event with another plant. For purposes of the present disclosure, the “event” refers to com event MON95275 or a modified corn event MON95275.

[0119] As used herein, the term “flanking” in reference to a transgenic event refers to the plant genomic sequence(s) immediately adjacent to the transgenic DNA insertion in the genome of a transformed plant, plant part, plant tissue, or plant cell comprising the transgenic event on the 5 ' and / or 3' side(s) or end(s) of the transgenic event insertion (e.g., corn event MON95275 or a modified corn event MON95275). Likewise, “flanking DNA”, “flanking sequence” or “flanking DNA sequence” each refers to a length of genomic DNA sequence immediately adjacent to the transgenic DNA insertion in the genome of the transformed plant on the 5' and / or 3' side(s) or end(s) of the insertion. A “5' flank” means the corn genomic DNA sequence adjacent to and upstream (or on the 5 ' end) of the transgenic DNA insertion. For example, a “5 ' flank” can include the com genomic DNA sequence immediately adjacent to and upstream (on the 5' end) of the transgenic insertion, or any com genomic DNA sequence upstream (on the 5 ' end) of the transgenic insertion that is not immediately adjacent to the transgenic insertion but is within about 5000 nucleotides, within about 4000 nucleotides, within about 3000 nucleotides, within about 2000 nucleotides, or within about 1000 nucleotides upstream of the transgenic insertion. Likewise, a “3' flank” means the corn genomic DNA sequence adjacent to and downstream (or on the 3' end)of the transgenic insert. For example, a “3' flank” can include the corn genomic DNA sequence immediately adjacent to and downstream (on the 3' end) of the transgenic insertion, or any com genomic DNA sequence downstream (on the 3 ' end) of the transgenic insertion that is not immediately adjacent to the transgenic insertion but is within about 5000 nucleotides, within about 4000 nucleotides, within about 3000 nucleotides, within about 2000 nucleotides, or within about 1000 nucleotides downstream of the transgenic insertion.

[0120] SEQ ID NOs: 11 and 12 are 1,073 and 1,106 nucleotide sequences representing corn (Zea mays) genomic DNA that flanks the transgenic insert at the 5' and 3' ends of the insert in corn event MON95275, respectively. SEQ ID NOs: 47 and 48 are 5,000 nucleotide sequences representing corn (Zea mays) genomic DNA that flanks the transgenic insert at the 5' and 3' ends of the insert, respectively. Nucleotides 3,928-5,000 of SEQ ID NO: 47 are identical to nucleotides 1-1,073 of SEQ ID NO: 11. Nucleotides 1-3,927 are based on the genomic sequence of the B73 corn cultivar (Zm-B73-REFERENCE-GRAMENE-4.0, NCBI). Similarly, nucleotides 1-1,106 of SEQ ID NO: 48 are identical to nucleotides 1-1,106 of SEQ ID NO: 12. The remaining nucleotides (1,107-5,000) are based on the genomic sequence of the B73 com cultivar.

[0121] The present disclosure provides the original transformant plant and progeny of the transformant that include the transgenic DNA and event. Such progeny may be produced by a sexual cross or outcross between plants comprising the same transgenic DNA and event, or between a plant comprising the transgenic DNA and event with another plant, or by any other method known in the art including any cell or tissue culture method, wherein the progeny includes the transgenic DNA and event. Such other plant may be a transgenic plant comprising the same and / or a different transgene or may be a non-transgenic plant, and each parental plant in a cross or outcross may be the same or different germplasm or breeding line. Even after repeated back- crossing to a recurrent parent, the transgenic DNA and event is present in progeny of the cross at the same chromosomal location. Thus, a “transgenic plant” can be the original transformant plant regenerated from the transformed plant cell and comprising the transgenic DNA and event, or a progeny plant of the original transformant plant, which may be separated from the transformant by one or more generations, that retains the transgenic DNA and event at the same specific location and sequence context in the plant’s genome. The transformant or progeny plant may be homozygous or heterozygous for corn event MON95275 or a modified corn event MON95275. In addition, a “transgenic plant” can include a plant produced from a transformed plant cell or tissue,or from another transgenic plant or plant part, by or using cell or tissue culture methods known in the art. A “transgenic plant” may comprise a plant having a transgene or transgenic event (including a modified transgenic event) stably inserted in the genome of at least one cell of the plant (z.e., corn event MON95275 or a modified corn event MON95275 in at least one cell of the plant), and the plant may be chimeric or non-chimeric with respect to the transgene and / or event or modified event. A transgenic plant is chimeric with respect to a transgene, event, or modified event if not all cells of the plant comprise the transgene, event, or modified event.

[0122] As used herein, the term “recombinant” refers to a non-natural DNA, protein, or combination that would not normally be found in nature, such as a combination of DNA sequences, proteins that would not naturally occur together, and is the result of human intervention. A “recombinant DNA molecule” is a DNA molecule comprising a combination of DNA sequences that would not naturally occur together and is the result of human intervention. Two or more elements of such combination of DNA sequences may be operably linked to one another. For example, a recombinant DNA molecule may comprise a combination of at least two DNA sequences that are heterologous with respect to each other, such as a DNA molecule that comprises a coding or transcribable DNA sequence operably linked to a heterologous promoter and / or other regulatory expression element(s), and / or a plant genomic DNA sequence comprising all or part of a transgene and a heterologous and flanking genomic sequence(s) adjacent to the transgene, and / or a DNA molecule that is artificially synthesized and comprises a polynucleotide sequence that deviates from any polynucleotide sequence that would normally exist in nature. A recombinant DNA molecule may comprise all or part of a junction sequence of the genome of a plant and all or part of the transgene insertion into the genome of the plant, and / or may comprise a recombinant or heterologous DNA fragment of event MON95275 or a modified corn event MON95275. An example of a recombinant DNA molecule is a DNA molecule comprising at least one of SEQ ID NOs: 1-10. As used herein, a recombinant plant, plant part, plant cell or plant tissue is a plant, plant part, plant cell or plant tissue that would not normally exist in nature, is the result of human intervention, and contains a transgene incorporated into the genome of the plant, plant part, plant cell or plant tissue. As a result of such genomic insertion, the recombinant plant is something new and distinctly different from any related wild-type or naturally occurring plant, plant part, plant cell or plant tissue. An example of a recombinant plant, plant part, plant cell or plant tissue is acorn or maize plant, plant part, plant cell or plant tissue containing the event MON95275 or a modified corn event MON95275.

[0123] As used herein, the term “heterologous” in reference to a combination of two or more DNA sequences or elements means that the two or more DNA sequences or elements do not normally exist together as such combination in nature without human intervention. As used herein, the term “heterologous” in reference to a DNA molecule, construct or sequence in relation to a plant, microorganism, plant cell or plant genome means that the DNA molecule, construct or sequence does not exist in nature as part of such plant, microorganism, plant cell or plant genome, and / or does not exist in the same physical or genomic location, context or orientation as part of such plant, microorganism, plant cell or plant genome in nature, without human intervention.

[0124] The present disclosure provides DNA molecules and fragments and their corresponding DNA sequences. The terms “DNA” and “DNA molecule” as used herein refer to a deoxyribonucleic acid (DNA) molecule. A DNA molecule may be of genomic or synthetic origin and / or comprise a recombinant or heterologous DNA molecule or sequence. A DNA molecule may be described in reference to its 5' (upstream) end and 3' (downstream) end. As used herein, the term “DNA sequence” refers to the polynucleotide sequence of the DNA molecule - i.e., the sequence of consecutive nucleotides in the DNA molecule. As used herein in reference to nucleotides of a polynucleotide or DNA sequence or molecule, the terms “consecutive” and “contiguous” are interchangeable and synonymous and refer to the 5' to 3' order of nucleotides in a polynucleotide or DNA sequence, strand or molecule without any gap or interruption between them. By convention, DNA sequences of the disclosure and fragments thereof are disclosed with reference to the 5 ' to 3 ' direction of only one strand of the two, anti-parallel and complementary DNA strands of a DNA molecule. By implication and intent, the complementary sequences of the sequences provided here (i.e., the sequences of the complementary, opposing, or antiparallel strand), also referred to in the art as the reverse complementary or reverse complement sequences, are within the scope of the present disclosure and are expressly intended to be within the potential scope of the subject matter as claimed. A DNA molecule, or a fragment derived therefrom, can also be extracted from plant part(s), plant cell(s) and / or tissue(s) or a homogenate, extract or lysate from plant part(s), plant cell(s) and / or tissue(s), or can be produced as an amplicon from extracted, purified or isolated DNA from plant part(s), plant cell(s) and / or tissue(s), or a homogenate, extractor lysate from plant part(s), plant cell(s) and / or tissue(s), which may further comprise event MON95275.

[0125] As used herein, the term “fragment” refers to a smaller piece or sequence of a larger or whole DNA molecule or sequence. For example, a fragment of SEQ ID NO: 9 or 10 may include a sequence that is at least about 12 consecutive nucleotides, at least about 13 consecutive nucleotides, at least about 14 consecutive nucleotides, at least about 15 consecutive nucleotides, at least about 16 consecutive nucleotides, at least about 17 consecutive nucleotides, at least about 18 consecutive nucleotides, at least about 19 consecutive nucleotides, at least about 20 consecutive nucleotides, at least about 21 consecutive nucleotides, at least about 22 consecutive nucleotides, at least about 23 consecutive nucleotides, at least about 24 consecutive nucleotides, at least about 25 consecutive nucleotides, at least about 30 consecutive nucleotides, at least about 35 consecutive nucleotides, at least about 40 consecutive nucleotides, at least about 45 consecutive nucleotides, at least about 50 consecutive nucleotides, at least about 60 consecutive nucleotides, at least about 70 consecutive nucleotides, at least about 80 consecutive nucleotides, at least about 90 consecutive nucleotides, at least about 100 consecutive nucleotides, at least about 200 consecutive nucleotides, at least about 300 consecutive nucleotides, at least about 400 consecutive nucleotides, or at least about 500 consecutive nucleotides of the larger, whole or complete DNA molecule or sequence of SEQ ID NO: 9 or 10.

[0126] According to present embodiments, a fragment of the DNA sequence of the 5' flank (SEQ ID NO: 11 or SEQ ID NO: 47) or the 3 ' flank (SEQ ID NO: 12 or SEQ ID NO: 48) of com event MON95275 can comprise at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, or at least 500 consecutive nucleotides of SEQ ID NO: 11 or SEQ ID NO: 47 or SEQ ID NO: 12 or SEQ ID NO: 48. It is possible that different maize germplasms may have differences in their genomic sequences, which may include differences in the flanking sequence(s), 5' flank and / or 3 ' flank of com event MON95275. These differences may result from introgression of the com event MON95275 or a modified corn event MON95275 into a different germplasm and / or spontaneous, mutagenic or genome editing changes that occur in a given germplasm or line. According to some embodiments, DNA molecules, constructs or polynucleotides are provided comprising a sequenceor flanking sequence, or a 5' flank or 3' flank, that is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8% or at least 99.9% identical to SEQ ID NO: 11 or 47 or SEQ ID NO: 12 or 48, or a fragment of either thereof. According to some embodiments, DNA molecules, constructs or polynucleotides are provided comprising a sequence or flanking sequence, or a 5' flank or 3 ' flank, that is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8% or at least 99.9% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, or at least 500 consecutive nucleotides of SEQ ID NO: 11 or 47 or SEQ ID NO: 12 or 48.

[0127] As used herein, the term “isolated” in reference to a molecule means that the molecule is at least partially separated from other molecules or sequences that are normally associated with the molecule in its native or natural state. In some embodiments, the term “isolated” refers to a DNA molecule that is at least partially separated from the nucleic acids or polynucleotide or DNA sequence(s) that normally flank and are covalently linked to the sequence of the DNA molecule in its native or natural state. An “isolated” DNA molecule may have a DNA sequence corresponding to a portion of the genome of a plant cell without other genomic DNA sequence(s) that normally flank and are covalently linked to the DNA sequence in nature. Such an “isolated” DNA molecule may comprise all or part of a transgene(s) and / or transgenic event, which may comprise all or part of corn event MON95275 or a modified corn event MON95275 or the transgene(s) or expression cassette(s) described herein. Nucleic acid sequences or elements, such as a coding sequence, intron sequence, untranslated leader sequence, promoter sequence, transcriptional termination sequence, and the like, that are naturally found within the DNA of the genome of an organism are not considered to be “isolated” so long as the element is within the genome of the organism and at the location within the genome in which it is naturally found. However, each of these elements, and subparts of these elements, would be “isolated” within the scope of this disclosure so long as the element or subpart is not within the genome of the organism, and at the location within the genomeof the organism, in which it is naturally found. An “isolated” DNA molecule may be any recombinant DNA molecule or amplification product or amplicon, and / or may comprise any DNA sequence removed from its natural or biological state and covalently fused to another DNA molecule or sequence with which it is not associated in nature. Such an isolated DNA molecule could be created by the use of biotechnology techniques, such as by making a recombinant DNA or integrating a foreign or heterologous DNA molecule into the chromosome of a cell, plant, or seed. Thus, any DNA molecule comprising a transgenic, recombinant, chimeric or artificial nucleotide sequence, transgene or expression cassette would be considered to be an “isolated” DNA molecule since these sequences are not naturally occurring, regardless of whether the sequence, transgene or expression cassette is present within a plasmid, vector or construct used to transform plant cells, within the genome of a plant, plant part, plant tissue, or plant cell, or is present in detectable amounts in tissues, progeny, biological samples or commodity products derived from a plant, plant part, plant tissue, or plant cell. A recombinant DNA molecule or sequence, or any fragment derived therefrom, comprising all or part of a transgene or junction sequence of com event would therefore also be considered to be “isolated.” An “isolated” DNA molecule may be extracted or purified from a transgenic plant(s), plant part(s), plant cell(s) and / or tissue(s), or may be present in a homogenate, extract or lysate from any such transgenic plant(s), plant part(s), plant cell(s) and / or tissue(s), or may be produced as an amplicon or amplification product from plant genomic DNA and / or extracted or purified DNA from transgenic plant(s), plant part(s), plant cell(s) and / or tissue(s), or a homogenate, extract or lysate from plant(s), plant part(s), plant cell(s) and / or tissue(s). For the purposes of this disclosure, any transgenic polynucleotide or DNA sequence, z.c., the nucleotide sequence of the DNA inserted into the genome of a plant or bacterium, or present in an extrachromosomal vector, would be considered to be an “isolated” nucleotide or DNA sequence whether it is present within the plasmid or similar structure used to transform the cells, within the genome of the plant or bacterium, or present in detectable amounts in tissues, progeny, biological samples or commodity products derived from the plant or bacterium. An “isolated” DNA molecule is a chemical or biochemical molecule, regardless of whether the molecule is referred to as a nucleic acid, a nucleic acid sequence, a polynucleotide sequence, a DNA sequence, a nucleic acid molecule, a polynucleotide molecule, a DNA molecule, or the like. An “isolated” molecule can provide industrial applicability when present in a plant cell or in aplant genome or when present outside of a plant cell, and therefore, provides and exhibits (and is intended to provide and exhibit) utility regardless of where the molecule is located.

[0128] The phosphodiester bond linkage between one end of a transgenic insert (or insertion) into the genome of a plant and the flanking com genomic DNA is referred to as a “junction.” In other words, a “junction” is the connection point or covalent linkage of one end of a transgenic insert and the flanking genomic DNA. One junction is found at the 5' end of the transgenic insertion and the other is found at the 3 ' end of the transgenic insert, referred to herein as the 5 ' and 3 ' junction, respectively. A “junction sequence” refers to a DNA sequence of any length of consecutive nucleotides that spans the 5' or 3' junction of a transgenic event in the plant genome. For a “junction sequence” to be specific to a junction between a transgenic event and a flanking genomic sequence, the junction sequence will generally comprise a sufficient number of consecutive nucleotides at one end of the insertion and a sufficient number of consecutive nucleotides of the flanking genomic sequence. According to some embodiments, a “junction sequence” may comprise (i) at least five (5) consecutive nucleotides, at least ten (10) consecutive nucleotides, at least fifteen (15) consecutive nucleotides, at least twenty (20) consecutive nucleotides, or at least thirty (30) consecutive nucleotides at one end of the insertion and (ii) at least five (5) consecutive nucleotides, at least ten (10) consecutive nucleotides, at least fifteen (15) consecutive nucleotides, at least twenty (20) consecutive nucleotides, or at least thirty (30) consecutive nucleotides of flanking genomic DNA sequence, although it is understood that any length of consecutive nucleotides spanning a junction of a transgenic event in a plant genome may be a junction sequence. A variety of junction sequences of corn event MON95275 can be determined by one of skill in the art using SEQ ID NO: 10. Examples of junction sequences of corn event MON95275 are provided as SEQ ID NOs: 1-8. FIG. 1 illustrates the physical arrangement and locations of the junction sequences, arranged from 5' to 3' (left to right), relative to SEQ ID NO: 10. The junction sequence(s) of a modified corn event MON95275 may be modified, mutated or edited relative to such junction sequence(s) of corn event MON95275. The junction sequences of corn event MON95275 or a modified corn event MON95275 may be present as part of the genome of a corn plant, plant part, plant seed, or plant tissue or cell containing com event MON95275 or a modified corn event MON95275, a DNA molecule containing all or part of corn event MON95275 or a modified com event MON95275, or a microorganism containing corn event MON95275 or a modified corn event MON95275. The identification of any one or more of the junction sequencesin a DNA molecule or sample from a plant, plant part, plant seed, or plant tissue or cell indicates that the plant, plant part, plant seed, or plant tissue or cell contains or comprises corn event MON95275 or a modified corn event MON95275, or the DNA molecule contains or comprises event MON95275 or a modified com event MON95275 or was obtained from a com plant, plant part, plant seed, or plant tissue or cell containing or comprising corn event MON95275 or a modified corn event MON95275, and is diagnostic in each case for the presence of corn event MON95275 or a modified corn event MON95275.

[0129] The junction sequences described herein can be diagnostic for the presence of all or part of corn event MON95275, or diagnostic for a modified corn event MON95275 if the junction sequence is unmodified in the modified com event MON95275, and / or a DNA molecule comprising all or part of the construct or expression cassettes described herein. Thus, the identification or detection, directly or indirectly, of one or more of SEQ ID NO: 1, SEQ ID NO:2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, and SEQ ID NO: 10 in a sample or DNA molecule derived from a corn plant, com plant part, corn seed, or corn tissue or cell, or a commodity product from a corn plant, corn plant part, com seed, or corn tissue or cell, is diagnostic that the corn plant, corn plant part, corn seed, or com tissue or cell, or a commodity product from a corn plant, corn plant part, com seed, or corn tissue or cell has or comprises all or part of corn event MON95275 or a modified corn event MON95275. The identification or detection, directly or indirectly, of a 5' junction sequence and a 3 ' junction sequence (each as provided or described herein) in a sample or DNA molecule derived from a corn plant, com plant part, com seed, or corn tissue or cell, or a commodity product from a corn plant, corn plant part, com seed, or corn tissue or cell, is diagnostic that the corn plant, com plant part, corn seed, or corn tissue or cell, or a commodity product from a corn plant, com plant part, com seed, or com tissue or cell has or comprises corn event MON95275 or a modified corn event MON95275. The present disclosure thus provides a DNA molecule that contains at least one of the nucleotide sequences provided as SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10. Any segment of DNA derived from transgenic com event MON95275 that is sufficient to include at least one of the sequences provided as SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO:3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10 is within the scope of the present disclosure. In addition, any DNA orpolynucleotide molecule or sequence comprising a sequence complementary to any of the sequences described herein is also within the scope of the present disclosure.

[0130] The disclosure provides DNA, polynucleotide or nucleic acid molecules, which may be single or double stranded, that can be used either as primers or probes for detecting the presence of DNA comprising all or part of event MON95275 or a modified corn event MON95275 in a sample derived from a corn plant, corn plant part, corn seed, or com tissue or cell, or a commodity product from a corn plant, corn plant part, corn seed, or com tissue or cell. Such primers or probes are specific for a target nucleic acid, polynucleotide or DNA sequence and, as such, are useful for the identification of corn event MON95275 nucleic acid, polynucleotide or DNA sequence, or a nucleic acid, polynucleotide, or DNA sequence of a modified com event MON95275, by the methods described herein. A primer or probe can hybridize to a target nucleic acid, polynucleotide or DNA sequence to allow for specific detection or amplification of a nucleic acid, polynucleotide or DNA molecule or sequence that comprises, or is covalently linked and associated with, the target nucleic acid, polynucleotide or DNA sequence. According to present embodiments, the primers and / or probe may be chosen to identify and distinguish detection of a particular transgenic event and not only the presence of a transgene in a plant genome. The target nucleic acid, polynucleotide or DNA molecule or sequence may comprise all or part of corn event MON95275 or a modified corn event MON95275, a junction sequence and / or flanking genomic DNA. Probes and primers according to the present disclosure may have (i) complete or 100% sequence complementarity (i.e., 100% complementary) to a target DNA sequence or (ii) incomplete sequence complementarity to a target DNA sequence, such as at least 60% complementary, at least 65% complementary, at least 70% complementary, at least 75% complementary, at least 80% complementary, at least 85% complementary, at least 90% complementary, at least 95% complementary, or at least 99% complementary to the target DNA sequence as long as the probe or primer has sufficient complementarity to the target DNA sequence to hybridize to the target DNA sequence under stringent hybridization conditions that are suitable and necessary for use of the probe or primer in the relevant amplification or detection assay, reaction or method. As understood in the art, the percentage complementarity of a primer or probe may be lower if the length of the primer or probe is longer and depends on the stringency and use.

[0131] A “probe” is a nucleic acid molecule that is complementary to a strand of target nucleic acid and is useful in hybridization methods. A probe may be attached a conventional detectablelabel or reporter molecule, e.g., a radioactive isotope, ligand, chemiluminescent agent, or enzyme. Such a probe is complementary to a strand of a target nucleic acid and, in the case of the present invention, to a strand of DNA from event MON95275 or a modified corn event MON95275, whether from an event MON95275 containing plant or from a plant containing a modified com event MON95275, or from a sample that includes event MON95275 DNA or DNA from a modified corn event MON95275. Probes according to the present invention include not only deoxyribonucleic or ribonucleic acids, but also polyamides and other probe materials that bind specifically to a target DNA sequence and can be used to detect the presence of that target DNA sequence. Examples of DNA sequences that may be useful as a probe for detecting corn event MON95275 or a modified corn event MON95275 are provided as: SEQ ID NO: 17 (PB10263), SEQ ID NO: 20 (PB50298), and SEQ ID NO: 23 (PRBNEG95275). A “probe” may also be used to bind a template DNA in a sample comprising all or part of a DNA or nucleotide sequence of corn event MON95275 or a modified corn event MON95275 to purify the template DNA from the remainder of the sample using purification methods or techniques known in the art, for example, if the probe is bound or can be bound to a substrate or a particle or bead that can be purified or separated. Such a template DNA may comprise all or part of a DNA or nucleotide sequence of corn event MON95275 or a modified corn event MON95275, or a portion or fragment thereof, such as a nucleotide sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10, or a complement thereof.

[0132] A “primer” is typically a DNA molecule that is designed for use in specific annealing or hybridization methods that involve thermal amplification. A pair of primers may be used with template DNA (such as a sample of corn genomic DNA) in a thermal amplification (such as polymerase chain reaction (PCR)) to produce an amplicon, where the amplicon produced from such reaction would have a DNA sequence corresponding to sequence of the template DNA located between the two sites where the primers hybridized to the template. As understood in the art, an “amplification product” or “amplicon” is a DNA molecule or segment produced by an amplification reaction. Amplification or amplifying refers to making multiple copies of a target DNA molecule or segment from a template DNA. A single “primer” may also be used to initiate a sequencing reaction to determine a DNA sequence of a template DNA according to sequencing methods known in the art. Such a sequencing reaction may be used to determine the presence orabsence of a DNA molecule or nucleotide sequence, or a portion or fragment thereof, from com event MON95275 or a modified corn event MON95275. Such a template DNA may comprise all or part of a DNA or nucleotide sequence of corn event MON95275 or a modified corn event MON95275, or a portion or fragment thereof, such as a nucleotide sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10, or a complement thereof.

[0133] DNA amplification reactions, methods and techniques are known to those skilled in art. DNA amplification can be accomplished by any of the various nucleic acid amplification methods known in the art, including thermal and isothermal amplification methods including the polymerase chain reaction or PCR. Amplification methods are known in the art and are described, inter alia, in U.S. Patent Nos. 4,683,195 and 4,683,202 and in PCR Protocols: A Guide to Methods and Applications, ed. Innis etal., Academic Press, San Diego, 1990. PCR amplification methods have been developed to amplify up to 22 kb (kilobase) of genomic DNA and up to 42 kb of bacteriophage DNA (Cheng et al., Proc. Natl. Acad. Sci. USA 91 :5695-5699, 1994). These methods as well as other methods known in the art of DNA amplification may be used in the practice of the present invention. Examples of DNA amplification methods include PCR, Recombinase Polymerase Amplification (RPA) (see for example U.S. Pat No. 7,485,428), Strand Displacement Amplification (SDA) (see for example, U.S. Pat. Nos. 5,455,166 and 5,470,723), Transcription-Mediated Amplification (TMA) (see for example, Guatelli et al., Proc. Natl. Acad. Sci. USA 87:1874-1878, 1990), Rolling Circle Amplification (RCA) (see for example, Fire and Xu, Proc. Natl. Acad Sci. USA 92:4641-4645, 1995; Lui, etal., J. Am. Chem. Soc. 118: 1587-1594, 1996; Lizardi, etal., Nature Genetics 19:225-232, 1998; U.S. Pat. Nos. 5,714,320 and 6,235,502), Helicase Dependent Amplification (HDA) (see for example Vincent et al., EMBO Reports 5(8): 795-800, 2004; U.S. Pat. No. 7,282,328), and Multiple Displacement Amplification (MDA) (see for example Dean et al., Proc. Natl. Acad Sci. USA 99:5261-5266, 2002). A sequence of the heterologous DNA insert and / or flanking genomic DNA sequence from corn event MON95275 can be verified or tested by amplifying such DNA molecules from corn seed containing event MON95275 DNA or corn plants grown from the corn seed containing event MON95275 DNA, using primers derived from the sequences provided herein, followed by standard DNA sequencing of the PCR amplicon or a cloned DNA fragment thereof.

[0134] According to present embodiments, the sequence of an amplicon of an amplification reaction may comprise one or more of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10, or a fragment thereof. According to present embodiments, the sequence of an amplicon comprises at least one junction sequence or two junction sequences, such as a 5 ' junction sequence and / or a 3' junction sequence for corn event MON95275 or a modified corn event MON95275.

[0135] A primer is typically designed to hybridize in a sequence-specific manner to a complementary target DNA strand to form a hybrid between the primer and target DNA strand, and the primer hybridized or bound to the complementary target DNA strand is a point of recognition for a polymerase to begin extension of the primer (z.e., polymerization of additional nucleotides into a lengthening nucleotide molecule) using as a template the target DNA strand. Primer pairs refer to use of two primers binding opposite strands of a double stranded DNA or polynucleotide segment for the purpose of amplifying the polynucleotide or DNA segment between the positions targeted for binding by the individual primers of the primer pair to the original template DNA or an amplicon of the amplification reaction, typically in a thermal cycling amplification reaction or other conventional DNA amplification method. Primer pairs are typically designed to hybridize to different nearby target positions of a template DNA molecule on opposing strands of the template DNA molecule such that the intervening region or sequence between the two primers can be specifically amplified for use or detection through multiple rounds of amplification. Exemplary DNA molecules useful as primers are provided as SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 18, SEQ ID NO: 19, SEQ ID NO: 21, SEQ ID NO: 22, SEQ ID NO: 24, SEQ ID NO: 25 and SEQ ID NO: 27.

[0136] The primer pair SEQ ID NO: 15 and SEQ ID NO: 16 are useful as a first DNA molecule and a second DNA molecule that is different from the first DNA molecule, and both are each of sufficient length of contiguous nucleotides of SEQ ID NO: 10 to function as DNA primers that, when used together in a thermal amplification reaction with template DNA derived from com event MON95275, to produce an amplicon diagnostic for corn event MON95275 DNA in a sample. The primer pair SEQ ID NO: 15 and SEQ ID NO: 16 are useful as a first DNA molecule and a second DNA molecule that is different from the first DNA molecule, and both are each of sufficient length of contiguous nucleotides of SEQ ID NO: 10 to function as DNA primers that, when used together in a thermal amplification reaction with template DNA derived from comevent MON95275, to produce an amplicon diagnostic for the zygosity of com event MON95275 DNA in a sample. The primer pair SEQ ID NO: 21 and SEQ ID NO: 22 are useful as a first DNA molecule and a second DNA molecule that is different from the first DNA molecule, and both are each of sufficient length of contiguous nucleotides of SEQ ID NO: 10 to function as DNA primers that, when used together in a thermal amplification reaction with template DNA derived from corn event MON95275, to produce an amplicon diagnostic for the zygosity of com event MON95275 DNA in a sample. The primer pair SEQ ID NO: 18 and SEQ ID NO: 19 are useful as a first DNA molecule and a second DNA molecule that is different from the first DNA molecule, and both are each of sufficient length of contiguous nucleotides of a locus within the corn genome to function as DNA primers that, when used together in a thermal amplification reaction with template DNA derived from corn event MON95275, to produce an amplicon that serves as an internal control for both the diagnosis of corn event MON95275, as well as the zygosity of com event MON95275 DNA in a sample.

[0137] According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises at least one of the nucleotide sequences provided as SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10. According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, or at least 90 consecutive nucleotides of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, or SEQ ID NO: 6. According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that (i) contains or comprises at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of any one of SEQ ID NOs: 1-10, (ii) is at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 nucleotides in length, and (iii) comprises nucleotides 1-1,001 and / or 3,733-3,734 of SEQ ID NO: 10.

[0138] According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises (i) two expression cassettes that encode the insect pesticidal insect toxins Cry75Aal, Vip4Da2, and the DvSnf7 specific dsRNA, and (ii) a junction sequence. According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises (i) expression cassettes that encodes the pesticidal insect toxins Cry75Aal and Vip4Da2, and the DvSnf7 specific dsRNA, and (ii) a 5' flank and / or a 3' flank sequence. The expression cassette may comprise in operable linkage: (a) a promoter sequence (b) a transcribable DNA sequence encoding a pesticidal insect toxins Cry75Aal and Vip4Da2, and the DvSnf7 specific dsRNA sequence that is toxic to Coleopteran insect pest species, and (c) a transcription termination or 3' UTR sequence. The expression cassette may further comprise any of the elements described in Table 1, which may be operably linked.

[0139] According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises (i) expression cassette that encodes Cry75Aal, Vip4Da2, and the DvSnf7 specific dsRNA, and (ii) a polynucleotide sequence comprising one or more of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, or SEQ ID NO: 10. According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises (i) expression cassettes that encodes the pesticidal insect toxins Cry75Aal, Vip4Da2, and the DvSnf7 specific dsRNA, and (ii) a polynucleotide sequence comprising at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of one or more of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, or SEQ ID NO: 10.

[0140] According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises (i) expression cassettes that encodes the pesticidal insect toxins Cry75Aal, Vip4Da2, and the DvSnf7 specific dsRNA, and (ii) a polynucleotide sequence comprising at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 1 1 or SEQ ID NO: 47 or SEQ ID NO: 12 or SEQ ID NO: 48. According to some embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises (i) expression cassettes that encodes the pesticidal insect toxins Cry75Aal, Vip4Da2, and the DvSnf7 specific dsRNA, and (ii) a sequence or flanking sequence, or a 5 ' flank or 3 ' flank, that is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99. 1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8% or at least 99.9% identical to SEQ ID NO: 11 or 47 or SEQ ID NO: 12 or 48, or a fragment of either thereof. According to some embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises (i) expression cassettes that an pesticidal insect toxins Cry75Aal, Vip4Da2, and the DvSnf7 specific dsRNA, and (ii) a sequence or flanking sequence, or a 5' flank or 3 ' flank, that is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99. 1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8% or at least 99.9% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 11 or 47 or SEQ ID NO: 12 or 48. According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises (i) expression cassettes that encodes the pesticidal insect toxins Cry75Aal, Vip4Da2, and the DvSnf7 specific dsRNA, and (ii) a polynucleotide sequence comprising one or morepolynucleotide sequences selected from SEQ ID NOs: 49-148. According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises (i) expression cassettes that encodes the pesticidal insect toxins Cry75Aal, Vip4Da2, and the DvSnf7 specific dsRNA, and (ii) a polynucleotide sequence comprising one or more polynucleotide sequences selected from SEQ ID NOs: 149-248.

[0141] According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises (i) expression cassettes that encodes the pesticidal insect toxins Cry75Aal, Vip4Da2, and the DvSnf7 specific dsRNA, (ii) a polynucleotide sequence comprising at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 11 or SEQ ID NO: 47, and (iii) a polynucleotide sequence comprising at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 12 or SEQ ID NO: 48. According to some embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises (i) expression cassettes that encodes the pesticidal insect toxins Cry75Aal, Vip4Da2, and the DvSnf7 specific dsRNA, (ii) a sequence or flanking sequence, or a 5' flank, that is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99. 1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8% or at least 99.9% identical to SEQ ID NO: 11 or 47, or a fragment of either thereof, and (iii) a sequence or flanking sequence, or a 3 ' flank, that is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8% or at least 99.9% identical to SEQ ID NO: 12 or 48, or a fragment of either thereof. According to some embodiments, a DNA molecule, construct, segment, amplicon, fragment orpolynucleotide is provided that contains or comprises (i) expression cassettes that encodes the pesticidal insect toxins Cry75Aal, Vip4Da2, and the DvSnf7 specific dsRNA, and (ii) a sequence or flanking sequence, or a 5' flank, that is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8% or at least 99.9% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 11 or 47, and (iii) a sequence or flanking sequence, or a 3 ' flank, that is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8% or at least 99.9% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 12 or 48. According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises (i) expression cassettes that encodes the pesticidal insect toxins Cry75Aal, Vip4Da2, and the DvSnf7 specific dsRNA, (ii) a polynucleotide sequence comprising one or more polynucleotide sequences selected from SEQ ID NOs: 49-148, and (iii) a polynucleotide sequence comprising one or more polynucleotide sequences selected from SEQ ID NOs: 149-248.

[0142] According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises expression cassettes that encodes the pesticidal insect toxins Cry75Aal, Vip4Da2, and the DvSnf7 specific dsRNA, wherein the DNA molecule, construct, segment, amplicon, fragment or polynucleotide comprises at its 5' end at least 50 consecutive nucleotides of SEQ ID NO: 11 or 47. According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide isprovided that contains or comprises expression cassettes that encodes the pesticidal insect toxins Cry75Aal, Vip4Da2, and the DvSnf7 specific dsRNA, wherein the DNA molecule, construct, segment, amplicon, fragment or polynucleotide comprises at its 3' end at least 50 consecutive nucleotides of SEQ ID NO: 12 or 48. According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises expression cassettes that encodes the pesticidal insect toxins Cry75Aal, Vip4Da2, and the DvSnf7 specific dsRNA, wherein the DNA molecule, construct, segment, amplicon, fragment or polynucleotide comprises at its 5' end a polynucleotide sequence selected from SEQ ID NOs: 49- 148. According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises expression cassettes that encodes the pesticidal insect toxins Cry75Aal, Vip4Da2, and the DvSnf7 specific dsRNA, wherein the DNA molecule, construct, segment, amplicon, fragment or polynucleotide comprises at its 3' end a polynucleotide sequence selected from SEQ ID NOs: 149-248.

[0143] According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises expression cassettes that encodes the pesticidal insect toxins Cry75Aal, Vip4Da2, and the DvSnf7 specific dsRNA, wherein the DNA molecule, construct, segment, amplicon, fragment or polynucleotide comprises at its 5' end at least 50 consecutive nucleotides of SEQ ID NO: 11 or 47, wherein the DNA molecule, construct, segment, amplicon, fragment or polynucleotide comprises at its 3 ' end at least 50 consecutive nucleotides of SEQ ID NO: 12 or 48, wherein the DNA molecule, construct, segment, amplicon, fragment or polynucleotide comprises at its 5' end a polynucleotide sequence selected from SEQ ID NOs: 49-148, and / or wherein the DNA molecule, construct, segment, amplicon, fragment or polynucleotide comprises at its 3' end a polynucleotide sequence selected from SEQ ID NOs: 149-248.

[0144] According to present embodiments, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 11 or SEQ ID NO: 47 at the 5' end of a DNA molecule, construct, segment, amplicon, fragment or polynucleotide may be immediately adjacent to and upstream (on the 5'end) of the transgenic insertion, or may not be immediately adjacent to, but further upstream (on the 5' end) and within about 5000 nucleotides, within about 4000 nucleotides, within about 3000 nucleotides, within about 2000 nucleotides, or within about 1000 nucleotides of the transgenic insertion. According to present embodiments, at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 12 or SEQ ID NO: 48 at the 3' end of a DNA molecule, construct, segment, amplicon, fragment or polynucleotide may be immediately adjacent to and downstream (on the 3' end) of the transgenic insertion, or may not be immediately adjacent to but further downstream (on the 3' end) and within about 5000 nucleotides, within about 4000 nucleotides, within about 3000 nucleotides, within about 2000 nucleotides, or within about 1000 nucleotides of the transgenic insertion. Any sequence comprising at least 50 consecutive nucleotides of SEQ ID NO: 11 or SEQ ID NO: 47, or at least 50 consecutive nucleotides of SEQ ID NO: 12 or SEQ ID NO: 48 is within the scope of the present disclosure. A DNA molecule, construct, segment, amplicon, fragment or polynucleotide can comprise at the 5' and / or 3' end of the construct (i) at least 50 consecutive nucleotides of SEQ ID NO: 11 or SEQ ID NO: 47; and / or (ii) at least 50 consecutive nucleotides of SEQ ID NO: 12 or SEQ ID NO: 48, respectively.

[0145] According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises (i) SEQ ID NO: 9 or a polynucleotide sequence comprising at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 9, and (ii) a junction sequence. According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises (i) SEQ ID NO: 9 or a polynucleotide sequence comprising at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, atleast 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 9, and (ii) a 5 ' flank and / or a 3 ' flank sequence.

[0146] According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises (i) SEQ ID NO: 9 or a polynucleotide sequence comprising at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 9, and (ii) a polynucleotide sequence comprising at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 11 or SEQ ID NO: 47 or SEQ ID NO: 12 or SEQ ID NO: 48. According to some embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises (i) SEQ ID NO: 9 or a polynucleotide sequence comprising at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 9, and (ii) a sequence or flanking sequence, or a 5' flank or 3 ' flank, that is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8% or at least 99.9% identical to SEQ ID NO: 11 or 47 or SEQ ID NO: 12 or 48, or a fragment of either thereof.

[0147] According to some embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises (i) SEQ ID NO: 9 or a polynucleotide sequence comprising at least 10, at least 11, at least 12, at least 13, at least 14, atleast 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21 , at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 9, and (ii) a sequence or flanking sequence, or a 5' flank or 3 ' flank, that is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8% or at least 99.9% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 11 or 47 or SEQ ID NO: 12 or 48. According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises (i) SEQ ID NO: 9 or a polynucleotide sequence comprising at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 9, and (ii) a polynucleotide sequence comprising one or more polynucleotide sequences selected from SEQ ID NOs: 49-148.

[0148] According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises (i) SEQ ID NO: 9 or a polynucleotide sequence comprising at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 9, and (ii) a polynucleotide sequence comprising one or more polynucleotide sequences selected from SEQ ID NOs: 149-248.

[0149] According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises (i) SEQ ID NO: 9 or a polynucleotide sequence comprising at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 9, (ii) a polynucleotide sequence comprising at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 11 or SEQ ID NO: 47, and (iii) a polynucleotide sequence comprising at least 10, at least 1 1, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 12 or SEQ ID NO: 48. According to some embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises (i) SEQ ID NO: 9 or a polynucleotide sequence comprising at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 9, (ii) a sequence or flanking sequence, or a 5' flank, that is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99. 1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8% or at least 99.9% identical to SEQ ID NO: 11 or 47, or a fragment of either thereof, and (iii) a sequence or flanking sequence, or a 3 ' flank, that is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%,at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8% or at least 99.9% identical to SEQ ID NO: 12 or 48, or a fragment of either thereof. According to some embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises (i) SEQ ID NO: 9 or a polynucleotide sequence comprising at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 9, and (ii) a sequence or flanking sequence, or a 5' flank, that is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8% or at least 99.9% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 11 or 47, and (iii) a sequence or flanking sequence, or a 3 ' flank, that is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99. 1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8% or at least 99.9% identical to at least 10, at least 1 1, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 12 or 48.

[0150] According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises (i) SEQ ID NO: 9 or a polynucleotide sequence comprising at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, atleast 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 9, (ii) a polynucleotide sequence comprising one or more polynucleotide sequences selected from SEQ ID NOs: 49-148, and (iii) a polynucleotide sequence comprising one or more polynucleotide sequences selected from SEQ ID NOs: 149-248.

[0151] According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises SEQ ID NO: 9 or a polynucleotide sequence comprising at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 9, wherein the DNA molecule, construct, segment, amplicon, fragment or polynucleotide comprises at its 5' end at least 50 consecutive nucleotides of SEQ ID NO: 11 or 47. According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises SEQ ID NO: 9 or a polynucleotide sequence comprising at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 9, wherein the DNA molecule, construct, segment, amplicon, fragment or polynucleotide comprises at its 3' end at least 50 consecutive nucleotides of SEQ ID NO: 12 or 48. According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises expression cassettes that encodes the pesticidal insect toxins Cry75Aal, Vip4Da2, and the DvSnf7 specific dsRNA, wherein the DNA molecule, construct, segment, amplicon, fragment or polynucleotide comprises at its 5 ' end a polynucleotide sequence selected from SEQ ID NOs: 49-148. According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises SEQ ID NO: 9 or a polynucleotide sequence comprising at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, atleast 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 9, wherein the DNA molecule, construct, segment, amplicon, fragment or polynucleotide comprises at its 3' end a polynucleotide sequence selected from SEQ ID NOs: 149-248.

[0152] According to present embodiments, a DNA molecule, construct, segment, amplicon, fragment or polynucleotide is provided that contains or comprises SEQ ID NO: 9 or a polynucleotide sequence comprising at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 9, wherein the DNA molecule, construct, segment, amplicon, fragment or polynucleotide comprises at its 5' end at least 50 consecutive nucleotides of SEQ ID NO: 11 or 47, wherein the DNA molecule, construct, segment, amplicon, fragment or polynucleotide comprises at its 3' end at least 50 consecutive nucleotides of SEQ ID NO: 12 or 48, wherein the DNA molecule, construct, segment, amplicon, fragment or polynucleotide comprises at its 5' end a polynucleotide sequence selected from SEQ ID NOs: 49-148, and / or wherein the DNA molecule, construct, segment, amplicon, fragment or polynucleotide comprises at its 3' end a polynucleotide sequence selected from SEQ ID NOs: 149-248.

[0153] According to present embodiments, a corn plant, plant part, plant seed, plant tissue, plant cell or commodity product is provided comprising any DNA molecule, construct, segment, fragment or polynucleotide described herein. A com plant comprising a construct as described herein may be further characterized as having resistance to Coleopteran insect pest species, relative to a non-transgenic control plant.

[0154] To detect the presence or absence of corn event MON95275 or a modified corn event MON95275, the target positions and / or the intervening region or sequence of a template DNA molecule may comprise at least one junction sequence and / or at least a portion of the insert of com event MON95275 or a modified com event MON95275. To detect the absence of corn event MON95275, the target positions and / or the intervening region or sequence of a template DNAmolecule may comprise corn genomic DNA that does not include a junction sequence or any portion of the insert of com event MON95275 or a modified com event MON95275. Thus, the presence or absence of an amplicon with a primer pair may be diagnostic of the presence or absence, respectively, of corn event MON95275 or a modified corn event MON95275 in a DNA molecule or sample, or vice versa. This may also be possible with more than one primer pair. For example, a first primer pair may produce a first amplicon if corn event MON95275 or a modified corn event MON95275 is present, and a second primer pair may produce a second amplicon if corn event MON95275 or a modified com event MON95275 is absent or not present. Alternatively, the size of an amplicon produced in an amplification reaction may also be diagnostic of the presence or absence of com event MON95275 or a modified com event MON95275 in a DNA molecule or sample - e.g., a primer pair may produce a first amplicon of a first size if corn event MON95275 or a modified corn event MON95275 is present or a second amplicon of a second size if corn event MON95275 or a modified corn event MON95275 is absent and not present; or a first primer pair may produce a first amplicon of a first size if corn event MON95275 or a modified com event MON95275 is present, and a second primer pair may produce a second amplicon of a second size if com event MON95275 or a modified corn event MON95275 is absent or not present. According to some of these embodiments, at least two primer pairs may be used wherein at least one of the primer pairs is used as an internal control and is not associated with corn event MON95275 or a modified corn event MON95275.

[0155] According to present embodiments, a primer pair to detect the presence of all or part of corn event MON95275 or a modified com event MON95275 in a DNA molecule or sample comprises a first primer and a second primer, wherein the first primer is complementary to a 5' flanking genomic DNA sequence and the second primer is complementary to a sequence within the transgenic insert; or wherein the first primer is complementary to a 5' flanking genomic DNA sequence and the second primer is complementary to a 3' flanking genomic DNA sequence; or wherein the first primer is complementary to a 3' flanking genomic DNA sequence and the second primer is complementary to a 5' flanking genomic DNA sequence; or wherein the first primer is complementary to a sequence within the transgenic insert and the second primer is complementary to a 3' flanking genomic DNA sequence; or wherein the first primer is complementary to a sequence within the transgenic insert and the second primer is complementary to a 5' flanking genomic DNA sequence; or wherein the first primer is complementary to a 3' flanking genomicDNA sequence and the second primer is complementary to a sequence within the transgenic insert. Each reference in this paragraph to a primer complementary to a 5' flanking genomic DNA sequence, a 3 ' flanking genomic DNA sequence, or a sequence within the transgenic insert of corn event MON95275 or a modified corn event MON95275 is also intended to potentially include a primer complementary to the reverse complement or opposing strand of the respective 5' flanking genomic DNA sequence, 3' flanking genomic DNA sequence, or sequence within the transgenic insert of corn event MON95275 or a modified corn event MON95275.

[0156] Examples of DNA molecules that may be useful as primers are provided as SEQ ID NO: 15, SEQ ID NO: 16, SEQ ID NO: 18, and SEQ ID NO: 19. The primer pair SEQ ID NO: 15 and SEQ ID NO: 16 can be useful as a first DNA molecule or primer and a second DNA molecule or primer, wherein each primer has sufficient length of consecutive nucleotides of SEQ ID NO: 10 or a sequence complementary to SEQ ID NO: 10 to function as DNA primers that, when used together in an amplification reaction with template DNA derived from corn event MON95275 or a modified corn event MON95275, hybridize to opposite strands of the template DNA and produce an amplicon diagnostic for corn event MON95275 DNA or DNA from a modified corn event MON95275 in a sample. The primer pair SEQ ID NO: 18 and SEQ ID NO: 19 are useful as a first DNA molecule or primer and a second DNA molecule or primer, wherein each primer has sufficient length of consecutive nucleotides of a locus within the corn genome to function as DNA primers that, when used together in a thermal amplification reaction with template DNA derived from com event MON95275 or a modified corn event MON95275, to produce an amplicon that serves as an internal control for both the diagnosis of com event MON95275 or a modified corn event MON95275, as well as the zygosity of corn event MON95275 DNA or DNA from a modified corn event MON95275 in a sample.

[0157] DNA probes and DNA primers are generally eleven (11) polynucleotides or more in length, often eighteen (18) polynucleotides or more, twenty -four (24) polynucleotides or more, or thirty (30) polynucleotides or more. Such probes and primers are selected to be of sufficient length to hybridize specifically to a target sequence under high stringency hybridization conditions. Preferably, probes and primers according to the present invention have complete sequence similarity with the target sequence, although probes differing from the target sequence that retain the ability to hybridize to target sequences may be designed by conventional methods.

[0158] The nucleic acid probes and primers of the present invention hybridize under stringent conditions to a target DNA molecule. Any conventional nucleic acid hybridization or amplification method can be used to identify the presence of DNA from a transgenic plant in a sample. Polynucleic acid molecules also referred to as nucleic acid segments or fragments thereof are capable of specifically hybridizing to other nucleic acid molecules under certain circumstances.

[0159] As used herein, two polynucleic acid molecules are said to be capable of specifically hybridizing to one another if the two molecules are capable of forming an anti-parallel, doublestranded nucleic acid structure. A nucleic acid molecule is said to be the “complement” of another nucleic acid molecule if they exhibit complete complementarity. As used herein, molecules are said to exhibit “complete complementarity” when every nucleotide of one of the molecules is complementary to a nucleotide of the other. Two molecules are said to be “minimally complementary” if they can hybridize to one another with sufficient stability to permit them to remain annealed to one another under at least conventional "low-stringency" conditions. Similarly, the molecules are said to be “complementary” if they can hybridize to one another with sufficient stability to permit them to remain annealed to one another under conventional "high-stringency" conditions. Conventional stringency conditions are described by Sambrook et al., 1989, and by Haymes etal., In: Nucleic Acid Hybridization, A Practical Approach, IRL Press, Washington, DC (1985). Departures from complete complementarity are therefore permissible, as long as such departures do not completely preclude the capacity of the molecules to form a double-stranded structure. In order for a nucleic acid molecule to serve as a primer or probe it need only be sufficiently complementary in sequence to be able to form a stable double-stranded structure under the particular solvent and salt concentrations employed.

[0160] As used herein, a substantially homologous sequence is a nucleic acid sequence that will specifically hybridize to the complement of the nucleic acid sequence to which it is being compared under high stringency conditions. Appropriate stringency conditions that promote DNA hybridization, for example, 6.0 x sodium chloride / sodium citrate (SSC) at about 45°C, followed by a wash of 2.0 x SSC at 50°C, are known to those skilled in the art or can be found in Current Protocols in Molecular Biology, John Wiley & Sons, N.Y. (1989), 6.3.1-6.3.6. For example, the salt concentration in the wash step can be selected from a low stringency of about 2.0 x SSC at 50°C to a high stringency of about 0.2 x SSC at 50°C. In addition, the temperature in the wash step can be increased from low stringency conditions at room temperature, about 22°C, to highstringency conditions at about 65°C. Both temperature and salt may be varied, or either the temperature or the salt concentration may be held constant while the other variable is changed. In a preferred embodiment, a polynucleic acid of the present invention will specifically hybridize to one or more of the nucleic acid molecules set forth in SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, or SEQ ID NO: 10, or complements thereof or fragments thereof under moderately stringent conditions, for example at about 2.0 x SSC and about 65°C. In a particularly preferred embodiment, a nucleic acid of the present invention will specifically hybridize to one or more of the nucleic acid molecules set forth in SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, or SEQ ID NO: 10, or complements or fragments thereof under high stringency conditions. In one aspect of the present invention, a preferred marker nucleic acid molecule of the present invention has the nucleic acid sequence set forth in SEQ ID NO: 1, or SEQ ID NO: 2, or SEQ ID NO: 3, or SEQ ID NO: 4, or SEQ ID NO: 5, or SEQ ID NO: 6, or SEQ ID NO: 7, or SEQ ID NO: 8, or SEQ ID NO: 9, or SEQ ID NO: 10, or complements thereof, or fragments thereof. The hybridization of the probe to the target DNA molecule can be detected by any number of methods known to those skilled in the art, these can include, but are not limited to, fluorescent tags, radioactive tags, antibody based tags, and chemiluminescent tags.

[0161] Regarding the amplification of a target nucleic acid sequence (e.g., by PCR) using a particular amplification primer pair, "stringent conditions" are conditions that permit the primer pair to hybridize only to the target nucleic acid sequence to which a primer having the corresponding wild-type sequence (or its complement) would bind and preferably to produce a unique amplification product, the amplicon, in a DNA thermal amplification reaction.

[0162] The term "specific for (a target sequence)" indicates that a probe or primer hybridizes under stringent hybridization conditions only to the target sequence in a sample comprising the target sequence.

[0163] As used herein, “amplified DNA” or “amplicon” refers to the product of polynucleic acid amplification method directed to a target polynucleic acid molecule that is part of a polynucleic acid template. For example, to determine whether a com plant resulting from a sexual cross contains transgenic plant genomic DNA from a corn plant comprising event MON95275 of the present invention, DNA that is extracted from a corn plant tissue sample may be subjected to apolynucleic acid amplification method using a primer pair that includes a first primer derived from a genomic DNA sequence in the region flanking the heterologous inserted DNA of event MON95275 and is elongated by polymerase 5 ' to 3' in the direction of the inserted DNA. The second primer is derived from the heterologous inserted DNA molecule is elongated by the polymerase 5' to 3' in the direction of the flanking genomic DNA from which the first primer is derived. The amplicon may range in length from the combined length of the primer pair plus one nucleotide base pair, or plus about fifty nucleotide base pairs, or plus about two hundred-fifty nucleotide base pairs, or plus about four hundred-fifty nucleotide base pairs or more. Alternatively, a primer pair can be derived from genomic sequence on both sides of the inserted heterologous DNA so as to produce an amplicon that includes the entire insert polynucleotide sequence (e.g., a forward primer isolated from the genomic portion on the 5' end of SEQ ID NO: 10 and a reverse primer isolated from the genomic portion on the 3' end of SEQ ID NO: 10 that amplifies a DNA molecule comprising the inserted DNA sequence (SEQ ID NO: 9) identified herein in the event MON95275 genome). A member of a primer pair derived from the plant genomic sequence adjacent to the inserted transgenic DNA is located a distance from the inserted DNA sequence, this distance can range from one nucleotide base pair up to about twenty thousand nucleotide base pairs. The use of the term “amplicon” specifically excludes primer dimers that may be formed in the DNA thermal amplification reaction.

[0164] For practical purposes, one should design primers which produce amplicons of a limited size range, for example, between 100 to 1000 bases. Smaller (shorter polynucleotide length) sized amplicons in general are more reliably produced in thermal amplification reactions, allow for shorter cycle times, and can be easily separated and visualized on agarose gels or adapted for use in endpoint TaqMan®-like assays. Smaller amplicons can be produced and detected by methods known in the art of DNA amplicon detection. In addition, amplicons produced using the primer pairs can be cloned into vectors, propagated, isolated, and sequenced or can be sequenced directly with methods well established in the art. Any primer pair derived from the combination of SEQ ID NO: 11 and SEQ ID NO: 9 or the combination of SEQ ID NO: 12 and SEQ ID NO: 9 that are useful in a DNA amplification method to produce an amplicon diagnostic for event MON95275 or a modified com event MON95275, or progeny thereof is an aspect of the present disclosure. Any single isolated DNA polynucleotide primer molecule comprising at least 15 contiguous nucleotides of SEQ ID NO: 11, or its complement that is useful in a DNA amplification methodto produce an amplicon diagnostic for event MON95275 or a modified com event MON95275 or progeny thereof is an aspect of the present disclosure. Any single isolated DNA polynucleotide primer molecule comprising at least 15 contiguous nucleotides of SEQ ID NO: 12, or its complement that is useful in a DNA amplification method to produce an amplicon diagnostic for plants comprising event MON95275 or a modified corn event MON95275 or progeny thereof is an aspect of the present disclosure. Any single isolated DNA polynucleotide primer molecule comprising at least 15 contiguous nucleotides of SEQ ID NO: 9, or its complement that is useful in a DNA amplification method to produce an amplicon diagnostic for event MON95275 or a modified corn event MON95275 or progeny thereof is an aspect of the present disclosure.

[0165] Polynucleic acid amplification can be accomplished by any of the various polynucleic acid amplification methods known in the art, including the polymerase chain reaction (PCR). Amplification methods are known in the art and are described, inter alia, in U.S. Patent Nos. 4,683,195 and 4,683,202 and in PCR Protocols: A Guide to Methods and Applications, ed. Innis et al., Academic Press, San Diego, 1990. PCR amplification methods have been developed to amplify up to 22 kb (kilobase) of genomic DNA and up to 42 kb of bacteriophage DNA (Cheng et al., Proc. Natl. Acad. Sci. USA 91:5695-5699, 1994). These methods as well as other methods known in the art of DNA amplification may be used in the practice of the present invention. The sequence of the heterologous DNA insert or flanking genomic DNA sequence from corn event MON95275 can be verified (and corrected if necessary) by amplifying such DNA molecules from corn seed containing event MON95275 DNA or corn plants grown from the corn seed containing event MON95275 DNA deposited with the ATCC having accession No. PTA- 126049, using primers derived from the sequences provided herein, followed by standard DNA sequencing of the PCR amplicon or cloned DNA fragments thereof.

[0166] A diagnostic amplicon produced by the methods described herein may be detected by a plurality of techniques known in the art, such as sequencing, restriction mapping, Northern analysis, Southern analysis, or any other suitable polynucleotide or DNA hybridization, blotting, polymerization and / or amplification based approach or technique. One method is Genetic Bit Analysis (Nikiforov et al., Nucleic Acid Res. 22:4167-4175, 1994) where a DNA oligonucleotide is designed that overlaps both the adjacent flanking genomic DNA sequence and the inserted DNA sequence - i.e., a junction sequence. The oligonucleotide is immobilized in wells of a microtiter plate. Following PCR of the region of interest (using, for example, one primer in the insertedsequence and one in the adjacent flanking genomic sequence), a single-stranded PCR product can be hybridized to the immobilized oligonucleotide and serve as a template for a single base extension reaction using a DNA polymerase and labeled dideoxynucleotide triphosphates (ddNTPs) specific for the expected next base. Readout may be fluorescent or ELISA-based. A signal indicates presence of the transgene / genomic junction sequence due to successful amplification, hybridization, and single base extension.

[0167] Another method is the Pyrosequencing technique as described by Winge (Innov. Pharma. Tech. 00: 18-24, 2000). In this method, an oligonucleotide is designed that overlaps the adjacent genomic DNA and insert DNA junction. The oligonucleotide is hybridized to single- stranded PCR product from the region of interest (one primer in the inserted sequence and one in the flanking genomic sequence) and incubated in the presence of a DNA polymerase, ATP, sulfurylase, luciferase, apyrase, adenosine 5' phosphosulfate and luciferin. dNTPs are added individually and the incorporation results in a light signal that is measured. A light signal indicates the presence of the transgene / genomic sequence due to successful amplification, hybridization, and single or multi-base extension.

[0168] Fluorescence Polarization as described by Chen, et al., (Genome Res. 9:492-498, 1999) is a method that can be used to detect the amplicon of the present invention. Using this method an oligonucleotide is designed that overlaps the genomic flanking and inserted DNA junction. The oligonucleotide is hybridized to single-stranded PCR product from the region of interest (one primer in the inserted DNA and one in the flanking genomic DNA sequence) and incubated in the presence of a DNA polymerase and a fluorescent-labeled ddNTP. Single base extension results in incorporation of the ddNTP. Incorporation can be measured as a change in polarization using a fluorometer. A change in polarization indicates the presence of the transgene / genomic sequence due to successful amplification, hybridization, and single base extension.

[0169] Real-time Polymerase Chain Reaction (PCR) is the ability to monitor the progress of the PCR as it occurs (i.e., in real time). Data is collected throughout the PCR process, rather than at the end of the PCR. In real-time PCR, reactions are characterized by the point in time during cycling when amplification of a target is first detected rather than the amount of target accumulated after a fixed number of cycles. In a real-time PCR assay, a positive reaction is detected by accumulation of a fluorescent signal. The higher the starting copy number of the nucleic acid target, the sooner a significant increase in fluorescence is observed. The cycle threshold (Ct value)is defined as the number of cycles required for the fluorescent signal to cross the threshold (z.e., exceeds background level). Ct levels are inversely proportional to the amount of target nucleic acid in the sample (i.e., the lower the Ct value, the greater the amount of target nucleic acid in the sample).

[0170] TaqMan® (PE Applied Biosystems, Foster City, CA) is described as a method of detecting and quantifying the presence of a DNA sequence using real-time PCR and is fully understood in the instructions provided by the manufacturer. Briefly, a FRET oligonucleotide probe is designed that overlaps the genomic flanking and insert DNA junction. The FRET probe and PCR primers (one primer in the insert DNA sequence and one in the flanking genomic sequence) are cycled in the presence of a thermalstable polymerase and dNTPs. Hybridization of the FRET probe results in cleavage and release of the fluorescent moiety away from the quenching moiety on the FRET probe. A fluorescent signal indicates the presence of the transgene / genomic sequence due to successful amplification and hybridization.

[0171] Other detection methods known in the art may be used. For example, microfluidics (see, e.g., U.S. Patent Publication No. 2006 / 068398; U.S. Patent No. 6,544,734) provide methods and devices that can be used to separate and amplify DNA samples or molecules. Optical dyes can be used to detect and measure specific DNA molecules (see, e.g., WO / 05017181). Nanotube devices (see, e.g., W 0 / 06024023) that comprise an electronic sensor for the detection of DNA molecules or nanobeads that bind specific DNA molecules can then be detected.

[0172] DNA detection kits that are based on DNA amplification methods contain DNA primer molecules that hybridize specifically to a target DNA and amplify a diagnostic amplicon under the appropriate reaction conditions. The kit may provide an agarose gel based detection method or any number of methods of detecting the diagnostic amplicon that are known in the art. DNA detection kits can be developed using the compositions disclosed herein and are useful for identification of corn event MON95275 DNA or DNA from a modified corn event MON95275 in a sample and can be applied to methods for breeding corn plants containing event MON95275 DNA or DNA from a modified com event MON95275. A kit that contains DNA primers that are homologous or complementary to any portion of the com genomic region as set forth in SEQ ID NO: 10 and to any portion of the inserted transgenic DNA as set forth in SEQ ID NO: 10 is an object of the invention. The DNA molecules can be used in DNA amplification methods (PCR) or as probes in polynucleic acid hybridization methods, i.e., southern analysis, northern analysis. Kits of theinvention may optionally also comprise reagents or instructions for performing the detection or diagnostic reactions described herein.

[0173] Probes and primers as provided herein may have complete sequence identity with the target sequence, although primers and probes differing from the target sequence that retain the ability to hybridize preferentially to target sequences may be designed by conventional methods. In order for a nucleic acid molecule to serve as a primer or probe it need only be sufficiently complementary in sequence to be able to form a stable double-stranded structure under the particular solvent and salt concentrations employed. Any conventional nucleic acid hybridization or amplification method can be used to identify the presence of transgenic DNA from corn event MON95275 or a modified corn event MON95275 in a sample.

[0174] Any number of methods well known to those skilled in the art can be used to isolate and manipulate a DNA molecule, or fragment thereof, disclosed herein, including DNA isolation or thermal amplification or PCR methods. Such DNA molecule or fragment may be inserted or placed into any suitable vector or plasmid or combined with other elements, sequences or fragments using molecular or recombinant techniques.

[0175] The DNA molecules and corresponding nucleotide sequences provided herein are therefore useful for, among other things, identifying corn event MON95275 or a modified corn event MON95275, detecting the presence of DNA derived from the transgenic corn event MON95275 or a modified corn event MON95275 in a sample, and monitoring samples for the presence and / or absence of corn event MON95275 or a modified corn event MON95275 or plant parts derived from corn plants comprising event MON95275 or a modified corn event MON95275.

[0176] Reference herein to “corn” generally is intended to include corn plants, corn plant cells, corn plant tissues, com seeds, corn plant parts, com progeny plants, and / or corn commodity products, depending on the context of its use herein, unless otherwise provided. The present disclosure provides corn plants, corn plant cells, corn plant tissues, corn seeds, corn plant parts (such as pollen, ovule, silk, spike, anther, cob, root tissue, stalk tissue, leaf tissue), corn progeny plants, and corn commodity products. These com plants, com plant cells, corn plant tissues, com seeds, corn plant parts, com progeny plants, and com commodity products contain a detectable amount of a polynucleotide or DNA molecule or sequence comprising at least one junction sequence and / or heterologous insert sequence of corn event MON95275 or a modified corn event MON95275, such as a polynucleotide or DNA molecule or sequence having or comprising at leastone of the sequences provided as SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10.

[0177] The present disclosure provides corn plants, com plant cells, corn seeds, com plant parts (such as pollen, ovule, silk, spike, anther, cob, root tissue, stalk tissue, leaf tissue), corn progeny plants derived from a transgenic corn plant containing event MON95275 DNA or DNA from a modified com event MON95275. A representative sample of corn seed containing event MON95275 DNA has been deposited according to the Budapest Treaty with the American Type Culture Collection (ATCC®). The ATCC repository has assigned the Patent Deposit Designation PTA- 126049 to the seed containing event MON95275 DNA.

[0178] The present disclosure provides a microorganism comprising a DNA molecule having at least one sequence selected from SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10 present in its genome. A microorganism is intended to include any microscopic cell or organism, whether prokaryote or eukaryote or otherwise, that contains DNA within a genome or chromosome or an extra-chromosomal DNA structure, such as a plasmid or vector, in such microscopic cell. Microscopic cells or organisms include bacteria (prokaryotes) and cells corresponding to higher life forms (eukaryotes) which are beneath the visual range of the average human. An example of such a microorganism is a transgenic plant cell. Microorganisms, such as a plant cell of the invention, are useful in many industrial applications, including but not limited to: (i) use as research tool for scientific inquiry or industrial research; (ii) use in culture for producing endogenous or recombinant carbohydrate, lipid, nucleic acid, or protein products or small molecules that may be used for subsequent scientific research or as industrial products; and (iii) use with modern plant tissue culture techniques to produce transgenic plants or plant tissue cultures that may then be used for agricultural research or production. The production and use of microorganisms such as transgenic plant cells utilizes modem microbiological techniques and human intervention to produce a man-made, unique microorganism. In this process, recombinant DNA is inserted into a plant cell’s genome to create a transgenic plant cell that is separate and unique from naturally occurring plant cells. This transgenic plant cell can then be cultured much like bacteria and yeast cells using modem microbiology techniques and may exist in an undifferentiated, unicellular state. The transgenic plant cell’s new genetic composition andphenotype is a technical effect created by the integration of the heterologous DNA into the genome of the cell. Another aspect of the invention is a method of using a microorganism of the invention. Methods of using microorganisms of the invention, such as transgenic plant cells, include (i) methods of producing transgenic cells by integrating recombinant DNA into the genome of the cell and then using this cell to derive additional cells possessing the same heterologous DNA; (ii) methods of culturing cells that contain recombinant DNA using modern microbiology techniques; (iii) methods of producing and purifying endogenous or recombinant carbohydrate, lipid, nucleic acid, or protein products from cultured cells; and (iv) methods of using modern plant tissue culture techniques with transgenic plant cells to produce transgenic plants or transgenic plant tissue cultures.

[0179] Corn plants of the present disclosure may pass along the event MON95275 DNA or DNA from a modified corn event MON95275, including transgene(s) or cassette(s) inserted in or part of corn event MON95275 or a modified corn event MON95275, to progeny or offspring. As used herein, “progeny” includes any plant, plant cell, seed, and / or regenerable plant part containing the event MON95275 DNA or DNA from a modified corn event MON95275 inherited or derived from an ancestor plant and / or comprising a DNA molecule having at least one sequence selected from SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10. Com plants, progeny, and seeds may be homozygous or heterozygous for the corn event MON95275 or a modified corn event MON95275 and / or the transgene(s) or cassette(s) of event MON95275 or a modified corn event MON95275. Progeny may be grown from seeds produced by a corn plant comprising or containing event MON95275 or a modified corn event MON95275 and / or from seeds produced by a plant fertilized with pollen from a corn plant comprising or containing event MON95275 or a modified com event MON95275 (i.e., fertilized with pollen comprising or containing corn event MON95275 or a modified corn event MON95275).

[0180] Methods for producing corn plants and seeds containing or comprising maize event MON95275 or a modified com event MON95275 are provided. Com plants may be bred using any method known in the art, for example, descriptions of breeding methods that are commonly used can be found in WR Fehr, in Breeding Methods for Cultivar Development, Wilcox J. ed., American Society of Agronomy, Madison WI (1987). Corn plants or progeny plants containing or comprising maize event MON95275 or a modified com event MON95275 may be self-pollinated(also known as “selfing”) to generate a true breeding line of com plants, z.e., corn plants homozygous for the transgene and event MON95275 or a modified corn event MON95275. Selfing can result in progeny known as an “inbred” that can be used to produce corn inbred lines that are genetically uniform.

[0181] Alternatively, corn plants or progeny plants containing or comprising maize event MON95275 or a modified corn event MON95275 may be out-crossed or cross-pollinated (also known as “crossing”), e.g., bred with another plant having a different germplasm or genotype, to produce a varietal or hybrid seed or plant that may be homozygous or heterozygous for the transgene(s) or cassette(s) and / or event MON95275 or a modified com event MON95275 depending on whether the other parental plant also comprises or contains the transgene(s) or cassette(s) and / or event MON95275 or a modified corn event MON95275. The other parental plant may be transgenic or non-transgenic for the same and / or different trait, transgene or event. A varietal or hybrid seed or plant of the invention may thus be derived by sexually crossing a first parent that lacks the specific and unique corn event MON95275 or a modified com event MON95275 with a second parent comprising corn event MON95275 or a modified corn event MON95275, resulting in a hybrid plant or progeny plant containing or comprising the specific and unique corn event MON95275 or a modified corn event MON95275. Each parent can be a hybrid or an inbred / varietal plant, so long as a parent or progeny plant or seed of the cross has or comprises at least one copy of the corn event MON95275 or a modified corn event MON95275 and / or a DNA molecule having or comprising at least one sequence selected from SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10.

[0182] Sexually crossing one plant with another plant, i.e., cross-pollinating, may be accomplished or facilitated by human intervention, for example: by human hands or other mechanical means under human, computer or automated control collecting the pollen of one plant and contacting this pollen with the style or stigma of a second plant; by human hands and / or human actions or other mechanical means under human, computer or automated control removing, destroying, devitalizing or covering the stamen or anthers of a plant (e.g., by manual intervention or by application of a chemical gametocide) so that natural self-pollination is prevented and crosspollination would have to take place in order for fertilization to occur; by human placement of pollinating insects in a position for “directed pollination” (e.g., by placing beehives in orchards orfields or by caging plants with pollinating insects); by human opening or removing of parts of the flower to allow for placement or contact of foreign pollen on the style or stigma; by selective placement of plants (e.g., intentionally planting plants in pollinating proximity); and / or by application of chemicals to precipitate flowering or to foster receptivity (of the stigma for pollen).

[0183] Two different transgenic plants may thus be crossed to produce hybrid offspring plants, plant parts and / or seeds that contain two independently segregating transgenes or events wherein at least one of those transgenes or events comprises or is contained within corn event MON95275 or a modified com event MON95275. For example, transgenic plants comprising corn event MON95275 or a modified corn event MON95275 can be crossed with other transgenic corn plants to produce a plant having the characteristics of both transgenic parents. Back-crossing to a parental plant and out-crossing with a non-transgenic plant are also contemplated, as is vegetative propagation. Descriptions of other breeding methods that are commonly used for different traits and crops are known in the art and can be found in one of several references, e.g., Fehr, in Breeding Methods for Cultivar Development, Wilcox J. ed., American Society of Agronomy, Madison WI (1987).

[0184] According to some embodiments progeny plants, plant parts or seeds may be analyzed using diagnostic methods as described herein to select for plants, plant parts or seeds containing or comprising com or maize event MON95275 or a modified corn event MON95275. Alternatively, progeny plants, plant parts or seeds may be analyzed using diagnostic methods as described herein to select for plants, plant parts or seeds containing or comprising corn or maize event MON95275 or a modified corn event MON95275.

[0185] Corn plants, progeny, seeds, cells and plant parts comprising corn event MON95275 or a modified com event MON95275, and / or one or transgene(s) or cassette(s) of corn event MON95275 or a modified corn event MON95275, may also contain one or more additional corn trait(s) or transgenic event(s), particularly those introduced by crossing a com plant containing such transgene(s) or cassette(s) and / or corn event MON95275 or a modified corn event MON95275 with another com plant containing the additional trait(s) or transgenic event(s). Such trait(s) or transgenic event(s) include, but are not limited to, increased insect resistance, herbicide tolerance, increased water use efficiency, increased yield performance, increased drought resistance, increased seed quality, improved nutritional quality, hybrid seed production, or disease or fungal resistance. A com trait may include any transgenic traits or mutant or edited traits oralleles. Mutant traits or alleles of a gene may be created by any mutagenesis technique known in the art, whereas edited traits may be generated by any genome editing technique or method known in the art. Many transgenic events in corn are known to those of skill in the art. For example, a list of such traits is provided by the United States Department of Agriculture’s (USDA) Animal and Plant Health Inspection Service (APHIS) and can be found on their website: www.aphis.usda.gov. Two or more trait(s) and / or transgenic event(s) comprising or including at least one copy of corn event MON95275 or a modified com event MON95275 may thus be combined in a progeny seed or plant by crossing two parent plants, each comprising one or more trait(s) or transgenic event(s), collecting the progeny seed, and selecting for progeny seed or plants that contain the two or more trait(s) and / or transgenic event(s). These steps may be repeated until the desired combination of trait(s) and / or transgenic event(s) in a progeny plant is achieved. For the present application, the progeny plant will generally comprise corn event MON95275 or a modified corn event MON95275. Back-crossing to a parental plant and out-crossing with a non- transgenic plant are also contemplated and is vegetative propagation.

[0186] A plant part is provided that comprises event MON95275 or a modified corn event MON95275 and / or is derived from corn plants comprising event MON95275 or a modified corn event MON95275. As used herein, a “plant part” refers to any part of a plant which may comprise event MON95275 or a modified com event MON95275 and / or material derived from a com plant comprising event MON95275 or a modified com event MON95275. Plant parts include, but are not limited to, plant tissue, pollen, ovule, silk, spike, anther, cob, root tissue, stalk tissue, and leaf tissue. Plant parts may be viable, nonviable, regenerable, and / or non-regenerable.

[0187] A commodity product is provided that is derived from one or more corn plants, plant parts, seeds and / or plant tissues comprising event MON95275 or a modified corn event MON95275 and that contains a detectable amount of a nucleic acid or DNA molecule, segment or sequence specific for event MON95275 or a modified com event MON95275. As used herein, a “commodity product” refers to any composition or product comprising material derived from one or more corn plants, whole or processed corn seed, one or more plant cells, and / or one or more plant parts containing or comprising the corn event MON95275 DNA or DNA from a modified com event MON95275. Nonviable commodity products include, but are not limited to, nonviable seeds, whole or processed seeds, seed parts, and plant parts; animal feed comprising corn, com oil, corn meal, corn flour, com flakes, com bran, pasta made with corn, com biomass, and fuel productsproduced using corn and com parts. Viable commodity products include, but are not limited to, seeds, plants, and plant cells. The corn plants comprising event MON95275 or a modified corn event MON95275 can thus be used to manufacture any commodity product typically acquired from com. Any such commodity product that is derived from corn plants comprising event MON95275 or a modified corn event MON95275 may contain at least a detectable amount of the specific and unique DNA corresponding to com event MON95275 or a modified corn event MON95275, and specifically may contain a detectable amount of a polynucleotide or DNA molecule having or comprising at least one sequence selected from SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10. Detection of one or more of these polynucleotide or DNA sequences in a sample may be used to determine or diagnose that the sample is taken from a com plant, corn plant part, corn plant tissue, corn plant cell, and / or corn plant product, such as a corn commodity product, comprising event MON95275 or a modified corn event MON95275, or to determine the content or source of a corn plant, com plant part, com plant tissue, com plant cell, and / or corn plant product, such as a corn commodity product. Any standard method of detection for nucleotide molecules may be used, including methods of detection disclosed herein. A commodity product is within the scope of the present disclosure if there is any detectable amount of a DNA molecule having at least one sequence selected from SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10 contained or comprised in the commodity product.

[0188] The corn plants, corn plant cells, corn seeds, corn plant parts (such as pollen, ovule, silk, spike, anther, cob, root tissue, stalk tissue, leaf tissue), corn progeny plants, and commodity products of the invention are therefore, useful for, among other things, growing plants for the purpose of producing seed and / or plant parts comprising corn event MON95275 or a modified corn event MON95275 for agricultural purposes, producing progeny comprising com event MON95275 or a modified com event MON95275 for plant breeding and research purposes, use with microbiological techniques for industrial and research applications, and sale to consumers.

[0189] Methods for producing an insect resistant corn plant comprising the DNA sequences specific and unique to event MON95275 or a modified corn event MON95275 of the present disclosure are provided. A progeny corn plant comprising the event MON95275 or a modified corn event MON95275 may be produced, for example, by selfing a parent plant or line comprisingthe event MON95275 or a modified com event MON95275, wherein such parent plant or line is homozygous or hemizygous for the event MON95275 or a modified corn event MON95275, or by crossing a first parent plant or line comprising the event MON95275 or a modified com event MON95275, wherein such parent plant or line is homozygous or hemizygous for the event MON95275 or a modified corn event MON95275, with a second parent plant or line having a different genotype or germplasm than the first parent line, wherein the second parent plant or line may or may not contain or comprise the event MON95275 or a modified corn event MON95275. As described further herein, a modified com event MON95275 may contain one or more of the expression cassette(s) or transgene(s) as provided herein, such as one or more of Cry75Aal, DvSnf7 dsRNA and / or Vip4Da2 expressing transgene cassette(s). According to some embodiments, the transgenic com plant(s) comprising the event MON95275 or a modified corn event MON95275 of the present disclosure may exhibit resistance to one or more Coleopteran insect pest species, such as Western Corn Rootworm (Diabrotica virgifera virgifera, WCR), and / or Northern Com Rootworm (Diabrotica barberi, NCR), relative to a non-transgenic control plant. Transgenic plants used in these methods may be homozygous or heterozygous for the transgene or event. Progeny plants produced by these methods may be varietal or hybrid plants; may be grown from seeds produced by plants containing com event MON95275 or a modified corn event MON95275 and / or from seeds produced by a plant fertilized with pollen from a plant containing corn event MON95275 or a modified com event MON95275; and may be homozygous or heterozygous for one or more transgene(s) described herein and / or com event MON95275 or a modified com event MON95275. Progeny plants may be subsequently self-pollinated to generate a true breeding line of plants, i.e., plants homozygous for the transgene, or alternatively may be out-crossed, e.g., bred with another unrelated plant, to produce a varietal or a hybrid seed or plant.

[0190] Methods of detecting the presence of DNA derived from a corn cell, corn tissue, com seed, or corn plant comprising corn event MON95275 or a modified com event MON95275 in a sample are provided. One method consists of (i) extracting a DNA sample from at least one com cell, corn tissue, com seed, or com plant; (ii) contacting the DNA sample with at least one primer that is capable of producing DNA sequence specific to event MON95275 DNA or DNA from a modified corn event MON95275 under conditions appropriate for DNA sequencing; (iii) performing a DNA sequencing reaction; and then (iv) confirming that the nucleotide sequence comprises a nucleotide sequence specific for event MON95275 or a modified corn eventMON95275, of the construct comprised therein, such as one selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10.

[0191] Another method consists of (i) extracting a DNA sample from at least one corn cell, corn tissue, corn seed, or com plant; (ii) contacting the DNA sample with a primer pair that is capable of producing an amplicon from event MON95275 DNA or DNA from a modified com event MON95275 under conditions appropriate for DNA amplification; (iii) performing a DNA amplification reaction; and then (iv) detecting the amplicon molecule and / or confirming that the nucleotide sequence of the amplicon comprises a nucleotide sequence specific for event MON95275 or a modified corn event MON95275, such as one selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10. The amplicon should be one that is specific for event MON95275, such as an amplicon that comprises SEQ ID NO: 1, or SEQ ID NO: 2, or SEQ ID NO: 3, or SEQ ID NO: 4, or SEQ ID NO: 5, or SEQ ID NO: 6, or SEQ ID NO: 7, or SEQ ID NO: 8, or SEQ ID NO: 9, or SEQ ID NO: 10. The detection of a nucleotide sequence specific for event MON95275 or a modified corn event MON95275 in the amplicon is determinative and / or diagnostic for the presence of the corn event MON95275 specific DNA or DNA specific for a modified corn event MON95275 in the sample. An example of a primer pair that is capable of producing an amplicon from event MON95275 DNA or DNA from a modified corn event MON95275 under conditions appropriate for DNA amplification is provided as SEQ ID NO: 15 and SEQ ID NO: 16. Other primer pairs may be readily designed by one of skill in the art and would produce an amplicon comprising SEQ ID NO: 1, or SEQ ID NO: 2, or SEQ ID NO: 3, or SEQ ID NO: 4, or SEQ ID NO: 5, or SEQ ID NO: 6, or SEQ ID NO: 7, or SEQ ID NO: 8 wherein such a primer pair comprises at least one primer within the genomic region flanking the insert and a second primer within the insert.

[0192] Another method of detecting the presence of DNA derived from a corn cell, com tissue, corn seed, or corn plant comprising corn event MON95275 or a modified corn event MON95275 in a sample consists of (i) extracting a DNA sample from at least one corn cell, corn tissue, corn seed, or corn plant; (ii) contacting the DNA sample with a DNA probe specific for event MON95275 or a modified com event MON95275; (iii) allowing the probe and the DNA sample to hybridize under stringent hybridization conditions; and then (iv) detecting hybridizationbetween the probe and the target DNA sample. An example of the sequence of a DNA probe that may be specific for event MON95275 or a modified corn event MON95275 is provided as SEQ ID NO: 17. Other probes may be readily designed by one of skill in the art and would comprise at least one fragment of genomic DNA flanking the insert and at least one fragment of insert DNA such as the sequence provided in SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, and SEQ ID NO: 10. Detection of probe hybridization to the DNA sample is diagnostic for the presence of com event MON95275 specific DNA or DNA specific for a modified corn event MON95275 in the sample. Absence of hybridization is alternatively diagnostic of the absence of corn event MON95275 specific DNA or DNA specific for a modified com event MON95275 in the sample.

[0193] DNA detection kits are provided that are useful for the identification of com event MON95275 DNA or DNA from a modified corn event MON95275 in a sample and can also be applied to methods for breeding corn plants containing the appropriate event DNA. Such kits may contain DNA primers and / or probe(s) which are specific for com event MON95275 or a modified corn event MON95275. Such DNA primers and / or probe(s) may comprise one or more of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10. One example of such a kit comprises at least one DNA molecule of sufficient length of continuous nucleotides of SEQ ID NO: 10 to function as a DNA probe useful for detecting the presence and / or absence of DNA derived from transgenic corn plants comprising event MON95275 or a modified corn event MON95275 in a sample. The DNA derived from transgenic corn plants comprising event MON95275 or a modified corn event MON95275 would comprise a DNA molecule having at least one sequence selected from SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10. The primers may comprise a primer pair including a first primer and a second primer, wherein at least one of the primers hybridizes to a flanking sequence and the other primer hybridizes to either an insert sequence of event MON95275 or a modified corn event MON95275 in the plant genome or the flanking sequence on the opposite side of the insert. The first and second primers hybridize to opposing strands of the com plant genomic DNA at different spaced apart positions such that an amplification reaction involving the two primers produces an amplicon comprising the primer sequences and the intervening sequence between the two primers. A probe may be chosen tocorrespond or hybridize to the amplicon produced with a primer pair or set of primers and may comprise all or part of the primer sequence(s) and / or the intervening sequence of the amplicon between the two primers. A DNA molecule that may be used as a DNA probe for determining, detecting, or diagnosing the presence and / or absence of corn event MON95275 or a modified com event MON95275 in a sample is provided as SEQ ID NO: 17. Suitable probes may be readily designed by one of skill in the art and should comprise at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 26, at least 27, at least 28, at least 29, at least 30, at least 31, at least 32, at least 33, at least 34, at least 35, at least 36, at least 37, at least 38, at least 39, or at least 40 contiguous nucleotides of SEQ ID NO: 10 and be sufficiently unique to com event MON95275 or a modified corn event MON95275 in order to identify DNA derived from the event.

[0194] Another type of kit comprises a primer pair useful for producing an amplicon useful for detecting the presence and / or absence of DNA derived from transgenic corn event MON95275 or a modified com event MON95275 in a sample. Such a kit would employ a method comprising contacting a target DNA sample with a primer pair as described herein, then performing a nucleic acid amplification reaction sufficient to produce an amplicon comprising a DNA molecule having at least one sequence selected from SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10 and then detecting the presence and / or absence of the amplicon. Such a method may also include sequencing the amplicon or a fragment thereof, which would be determinative of, i.e., diagnostic for, the presence of the corn event MON95275 specific DNA or DNA specific for a modified com event MON95275 in the target DNA sample. Other primer pairs may be readily designed by one of skill in the art and should comprise at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 26, at least 27, at least 28, at least 29, or at least 30 contiguous nucleotides of sequences provided in, but not limited to SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10, and be sufficiently unique to corn event MON95275 DNA or DNA from a modified corn event MON95275 in order to identify DNA derived from the event.

[0195] The kits and detection methods of the invention are useful for, among other things, identifying corn event MON95275 or a modified com event MON95275, selecting plant varietiesor hybrids comprising com event MON95275 or a modified corn event MON95275, detecting the presence of DNA derived from the transgenic com plant comprising event MON95275 or a modified corn event MON95275 in a sample, and monitoring samples for the presence and / or absence of com plants comprising event MON95275 or a modified corn event MON95275, or plant parts derived from corn plants comprising event MON95275 or a modified corn event MON95275.

[0196] The sequences of the heterologous DNA insert, junction sequences, or flanking sequence from com event MON95275 or a modified com event MON95275 can be verified (and corrected if necessary) by amplifying such sequences from the event using primers derived from the sequences provided herein followed by standard DNA sequencing of the amplicon or of the cloned DNA.

[0197] Methods of detecting the zygosity of the transgene allele of DNA derived from a corn cell, corn tissue, corn seed, or corn plant comprising corn event MON95275 or a modified corn event MON95275 in a sample are provided. One method consists of (i) extracting a DNA sample from at least one corn cell, com tissue, corn seed, or corn plant; (ii) contacting the DNA sample with a primer pair that is capable of producing a first amplicon diagnostic for event MON95275 or a modified com event MON95275; (iii) contacting the DNA sample with a primer pair that is capable of producing a second amplicon diagnostic for native com genomic DNA not comprising event MON95275; (iv) performing a DNA amplification reaction; and then (v) detecting the amplicons, wherein the presence of only the first amplicon is diagnostic of a homozygous event MON95275 DNA or DNA from a modified corn event MON95275 in the sample, and the presence of both the first amplicon and the second amplicon is diagnostic of a com plant heterozygous for event MON95275 or a modified com event MON95275. An example of a set of primer pairs are presented as SEQ ID NO: 15 and SEQ ID NO: 16 which produce an amplicon diagnostic for event MON95275 or a modified corn event MON95275; and SEQ ID NO: 21 and SEQ ID NO: 22 which produces an amplicon diagnostic for non-inserted wild-type corn genomic DNA not comprising event MON95275 or a modified com event MON95275. A set of probes can also be incorporated into such an amplification method to be used in a real-time PCR format using the primer pair sets described above. An exemplary set of probes are presented as SEQ ID NO: 17 (diagnostic for the amplicon for the event MON95275 or a modified com event MON95275) and SEQ ID NO: 23(diagnostic for the amplicon for wild-type corn genomic DNA not comprising event MON95275 or a modified corn event MON95275).

[0198] Another method for determining zygosity consists of (i) extracting a DNA sample from at least one corn cell, corn tissue, com seed, or corn plant; (ii) contacting the DNA sample with a probe set which contains at least a first probe that specifically hybridizes to event MON95275 DNA or DNA from a modified com event MON95275and at least a second probe that specifically hybridizes to com genomic DNA that was disrupted by insertion of the heterologous DNA of event MON95275 or a modified corn event MON95275 and does not hybridize to event MON95275 DNA or DNA from a modified com event MON95275; (iii) hybridizing the probe set with the sample under stringent hybridization conditions, wherein detecting hybridization of only the first probe under the hybridization conditions is diagnostic for a homozygous allele of event MON95275 DNA or DNA from a modified corn event MON95275 in the sample; and wherein detecting hybridization of both the first probe and the second probe under the hybridization conditions is diagnostic for a heterozygous allele of event MON95275 or a modified corn event MON95275 in a DNA sample; and wherein detecting hybridization of only the second probe under the hybridization conditions is diagnostic for the absence of corn event MON95275 DNA or DNA from a modified com event MON95275 in the sample.

[0199] Yet another method for determining zygosity consists of (i) extracting a DNA sample from at least one corn cell, com tissue, corn seed, or corn plant; (ii) contacting the DNA sample with a primer pair that is capable of producing an amplicon from one or more of the toxin coding sequences encoding Cry75Aal, Vip4Da2, and / or the DvSnf7 specific dsRNA and / or DNA from corn event MON95275 or a modified corn event MON95275; (iii) contacting the DNA sample with a primer pair that is capable of producing an amplicon of an internal standard known to be single-copy and homozygous in the com plant; (iv) contacting the DNA sample with a probe set which contains at least a first probe that specifically hybridizes to one or more of the toxin coding sequences encoding Cry75Aal, Vip4Da2, and / or the DvSnf7 specific dsRNA and / or DNA from corn event MON95275 or a modified corn event MON95275, and at least a second probe that specifically hybridizes to the internal standard genomic DNA known to be single-copy and homozygous in the com plant; (v) performing a DNA amplification reaction using real-time PCR and determining the cycle thresholds (Ct values) of the amplicon corresponding to the toxin coding sequence and the single-copy, homozygous internal standard; (vi) calculating the difference (ACt)between the Ct value of the single-copy, homozygous internal standard amplicon and the Ct value of the toxin coding sequence amplicon; and (vii) determining zygosity, wherein a ACt of around zero (0) indicates homozygosity of the inserted T-DNA and a ACt of around one (1) indicates heterozygosity of the inserted T-DNA. Heterozygous and homozygous events are differentiated by a ACt value unit of approximately one (1). Given the normal variability observed in real-time PCR due to multiple factors such as amplification efficiency and ideal annealing temperatures, the range of “about one (1)” is defined as a ACt of 0.75 to 1.25. Primer pairs and probes for the above method for determining zygosity can amplify and detect amplicons from the Cry75Aal cassette or coding sequence, the Vip4Da2 cassette or coding sequence, the DvSnf7 specific dsRNA cassette or coding sequence, and / or a junction sequence. An exemplary primer pair for the detection of the amplicon corresponding to the 3 ' junction sequence are presented as SEQ ID NO: 15 combined with SEQ ID NO: 16 and the internal standard presented as SEQ ID NO: 18 combined with SEQ ID NO: 19. The accompanying exemplary probes are presented as SEQ ID NO: 17 (3' junction sequence) and SEQ ID NO: 20 (internal standard).

[0200] According to embodiments of the present disclosure, a transgenic corn plant or plant part, one or more transgenic corn plants or plant parts or a plurality transgenic com plants or plant parts as provided herein, or an agricultural field or soil in which a transgenic com plant or plant part, one or more transgenic corn plants or plant parts or a plurality of transgenic com plants or plant parts as provided herein are planted or grown, can be treated with an agricultural composition comprising one or more active ingredients or other agents, such as, for example and without limitation, an herbicide or one or more herbicides, a fungicide or one or more fungicides, an insecticide or one or more insecticides, a plant growth regulator or plant stimulant or one or more plant growth regulators and / or plant stimulants, and / or a safener or one or more safeners. Provided below are lists of possible or representative compounds for each of these types of actives or agents, and an agricultural composition may comprise one or any combination or multiplicity of these actives, agents or compounds. Such an agricultural composition may be applied, for example, as a foliar, soil or in-furrow treatment, as a pre-emergent, pre-sowing and / or post-emergent treatment, and / or in some cases, may be applied to a transgenic plant part or seed provided herein.

[0201] An agricultural composition may be formulated according to its intended use and application. The appropriate formulation of the agricultural composition may be chosen to have different physicochemical parameters, components and stabilities of the respective compound(s).Possible types of formulations for an agricultural composition can include, for example: wettable powders (WP), water-soluble powders (SP), water-soluble concentrates, emulsifiable concentrates (EC), emulsions (EW), such as oil-in-water and water-in-oil emulsions, sprayable solutions, suspension concentrates (SC), dispersions based on oil or water, oil-miscible solutions, capsule suspensions (CS), dusting products (DP), dressings, granules for scattering and soil application, granules (GR) in the form of microgranules, spray granules, absorption and adsorption granules, water-dispersible granules (WG), water-soluble granules (SG), ULV formulations, microcapsules and waxes. If appropriate, some agricultural compositions of a pesticidal compound or one or more pesticidal compounds might be formulated and used as a seed coating applied to a plant part or seed as provided herein.

[0202] Plants, progeny, plant parts, plant seeds, plant tissues, and plant cells may contain or comprise one or more additional desirable trait(s). Such desirable traits may be transgenic traits, native traits, or traits produced by other methods, such as genome editing, base editing, prime editing or other conventional mutagenesis methods. Such desirable trait(s) may provide an agronomic, agricultural or commodity benefit to a plant, plant part, plant seed or plant product. Desirable traits may be combined with com event MON95275 or a modified corn event MON95275 by, for example, crossing a corn plant comprising corn event MON95275 or a modified com event MON95275 with another corn plant containing the additional trait(s). Alternatively, a trait may be created by mutagenesis, editing or site-directed integration of or into a plant, plant part or plant cell comprising corn event MON95275 or a modified corn event MON95275. Such traits may include, but are not limited to, increased insect resistance, increased water use efficiency, increased nitrogen use efficiency, increased yield performance, increased drought resistance, increased disease resistance, increased seed quality, improved nutritional quality, hybrid seed production, and / or increase herbicide tolerance, in which the trait is measured with respect to a com plant lacking such transgenic trait. For example, the MON95275 event or a modified corn event MON95275 could be stacked by breeding or introgression with another event(s), or a combination of events, known in the art including, but not limited to:• MON00603 (also known as NK603 or MON603; Roundup Ready™ 2 Maize for herbicide tolerance; deposited as ATCC PTA-2478 and described in US Patent Application Publication No. 2007 / 292854 and US Patent No. 6,825,400, the entire contents and disclosure of each of which are incorporated herein by reference),• MON89034 (YieldGard™ VT Pro™ for insect resistance; deposited as ATCC PTA-7455 and described in PCT Publication No. W02007 / 140256 and US Patent Application Publication No. US2008 / 260932, the entire contents and disclosure of each of which are incorporated herein by reference),• MON88017 (YieldGard™ VT™ Rootworm™ RR2 for herbicide tolerance and insect resistance; deposited as PTA-5582 and described in US Patent Application Publication No. 2008 / 028482 and PCT Publication No. W02005 / 059103, the entire contents and disclosure of each of which are incorporated herein by reference),• MON87427 (Roundup Ready™ Maize for herbicide tolerance, deposited as ATCC PTA- 7899, described in US Patent No. 8,618,358 and PCT Publication No. WO2011 / 062904, the entire contents and disclosure of each of which are incorporated herein by reference),• MON87411 (for insect resistance; deposited as ATCC No. PTA-12669 and described in US Patent No. 10,316,330 and PCT Publication No. WO2013 / 169923, the entire contents and disclosure of each of which are incorporated herein by reference),• MON87429 (for herbicide tolerance; deposited as ATCC PTA-124635 and described in US Patent No. 10,920,239 and PCT Publication No. WO2019 / 152316, the entire contents and disclosure of each of which are incorporated herein by reference),• MON87460 (Genuity® DroughtGard™ for abiotic stress tolerance; deposited as ATCC No. PTA-8910 and described in PCT Publication No. W02009 / 111263 and US Patent Application Publication No. 2011 / 0138504, the entire contents and disclosure of each of which are incorporated herein by reference),• MON87419 (for herbicide tolerance; deposited as ATCC PTA-120860 and described in US Patent No. 11,098,321 and PCT Publication WO2015 / 142571, the entire contents and disclosure of each of which are incorporated herein by reference),• MON95379 (for insect resistance; deposited as ATCC PTA-125027 and described in PCT Publication W02020 / 028172) and US Patent Application Publication No. US2020 / 032289, the entire contents and disclosure of each of which are incorporated herein by reference.• MGN00810 (for insect resistance; also known as MON810; described in US Patent Application Publication No. 2002 / 102582, the entire contents and disclosure of which are incorporated herein by reference),• MON00021 (also known as GA21; Roundup Ready™ Maize, Agrisure™GT for herbicide tolerance, deposited as ATCC 209033 and described in US Patent Application Publication No. 2005 / 086719 and PCT Publication No. WO1998 / 044140, the entire contents and disclosure of each of which are incorporated herein by reference),• MON832 (Roundup Ready™ Maize for herbicide tolerance),• MON863 (YieldGard™ Rootworm RW, MaxGard™ for insect resistance; deposited as ATCC PTA-2605 and described in PCT Publication No. W02004 / 011601 and US Patent Application Publication No. 2006 / 095986, the entire contents and disclosure of each of which are incorporated herein by reference),• AGV-PY203-4 (GralNzyme Phytase for modified product quality),• ACS-ZM004-3 (Starlink™ Maize for herbicide tolerance and insect resistance),• ACS-ZM001-9 (InVigor™ Maize for pollination control system),• ACS-ZM005-4 (InVigor™ Maize for pollination control system),• ACS-ZM002-1 (Liberty Link™ Maize for herbicide tolerance),• ACS-ZM003-2 (Liberty Link™ Maize for herbicide tolerance),• DAS-40278-9 (Enlist™ Maize for herbicide tolerance, deposited as ATCC No. PTA- 10244 and described in US Patent No. 11,098,322 and PCT Publication No. WO20 11 / 022469),• DAS-01507-1 (also known as TC1507; Herculex™ I, Herculex™ CB for herbicide tolerance and insect resistance; described in US Patent Application Publication No. 2005039226 and PCT Publication No W02004 / 099447),• DAS-59122-7 (Herculex™ RW for herbicide tolerance and insect resistance; described in US Patent Application Publication No. 2006 / 070139),• DKB-89614-9 (Bt Xtra™ Maize for herbicide tolerance and insect resistance),• DP-32138-1 (32138 SPT maintainer for pollination control system; deposited as ATCC No. PTA-9158 and described in US Patent Application Publication No. 2009 / 0210970 and PCT Publication No. W02009 / 103049),• DP-098140-6 (Optimum™ GAT™ for herbicide tolerance; deposited as ATCC No. PTA- 8296 and described in US Patent Application Publication No. 2009 / 137395 and PCT Publication No. W02008 / 112019),• MIR162 (Agrisure™ Viptera for insect resistance; deposited as ATCC No. PTA-6188 and described in US Patent Application Publication No. 2009 / 300784 and PCT Publication No. W02007 / 142840),• MIR604 (Agrisure™ RW for insect resistance, described in US Patent Application Publication No. 2008 / 167456 and PCT Publication No. W02005 / 103301),• REN-00038-3 (also known as LY038; Mavera™ Maize for modified product quality; deposited as ATCC No. PTA-5623 and described in PCT Publication No. W02005 / 061720 and US Patent No. 7,615,621, the entire contents and disclosure of each of which are incorporated herein by reference),• SYN-E3272-5 (Enogen™ for modified product quality, described in US Patent No. 7,635,799 and PCT Application Publication No. W02006 / 098952),• SYN-05307-1 (Agrisure® Duracade™ for insect resistance; deposited as ATCC No. PTA- 9561 and described in PCT Publication No. W02010 / 077816 and US Patent No. US10,100,371),• BtlO (for herbicide tolerance and insect resistance),• SYN-BT011-1 (Agrisure™ CB / LL for herbicide tolerance and insect resistance),• SYN-EV176-9 (NaturGard KnockOut™, Maximizer™ for herbicide tolerance and insect resistance),• MON89034 x DAS-01507-1 x MON603 x MIR162 x DAS-40278-9 (Power Core™ x MIR162 x Enlist™ for herbicide tolerance and insect resistance),• DAS-01507-1 x DAS-59122-7 (Herculex XTRA™ for herbicide tolerance and insect resistance),• DAS-01507-1 x DAS-59122-7 x MON603 (Herculex XTRA™ RR for herbicide tolerance and insect resistance),• DAS-01507-1 x MON603 (Herculex™ I RR for herbicide tolerance and insect resistance),• DAS-59122-7 x MON603 (Herculex™ RW Roundup Ready™ 2 for herbicide tolerance and insect resistance),• DAS-01507-1 x DAS-59122-7 x MDN00810 x MIR604 x MON603 (Optimum™ Intrasect Xtreme for herbicide tolerance and insect resistance),• DAS-01507-1 x DAS-59122-7 x MON810 x MON603 (Optimum™ Intrasect XTRA for herbicide tolerance and insect resistance),• DAS-01507-1 x MIR604 x MON603 (Optimum™ TRIsect for herbicide tolerance and insect resistance),• DAS-01507-1 x MON810 x MON603 (Optimum™ Intrasect for herbicide tolerance and insect resistance),• MON00021 x MON810 (Roundup Ready™ YieldGard™ Maize for herbicide tolerance and insect resistance),• MON810 x MON88017 (YieldGard™ VT Triple for herbicide tolerance and insect resistance),• MON863 x MON810 (YieldGard™ Plus for insect resistance),• MON603 x MON810 x MON863 (YieldGard™ Plus with RR for herbicide tolerance and insect resistance),• MON863 x MON603 (YieldGard™ RW + RR for herbicide tolerance and insect resistance),• MON87427 x MON89034 x DAS-01507-1 x MON87411 x DAS-59122-7 x DAS-40278-9 (SmartStax™ Pro x Enlist™ for herbicide tolerance and insect resistance),• MON89034 x MON88017 (Genuity® VT Triple Pro™ for herbicide tolerance and insect resistance),• MON89034 x MON603 (Genuity® VT Double Pro™ for herbicide tolerance and insect resistance),• MON89034 x DAS-01507-1 x MON88017 x DAS-59122-7 (Genuity® SmartStax™ for herbicide tolerance and insect resistance),• MON89034 x DAS-01507-1 x MON603 (Power Core™ for herbicide tolerance and insect resistance),• MON603 x M0N810 (YieldGard™ CB + RR for herbicide tolerance and insect resistance),• MON603 x ACS-ZM003-2 (Roundup Ready™ Liberty Link™ Maize for herbicide tolerance),• ACS-ZM003-2 x M0N810 (Liberty Link™ Yieldgard™ Maize for herbicide tolerance and insect resistance),• REN-00038-3 x MON810 (Mavera™ YieldGard™ Maize for insect resistance and modified product quality),• SYN-05307-1 X MIR604 x SYN-BT011-1 x DAS-01507-1 x MQN00021 (Agrisure® Duracade™ 5122 for herbicide tolerance and insect resistance),• SYN-05307-1 x MIR604 x SYN-BT011-1 x DAS-01507-1 x MON00021 x MIR162 (Agrisure® Duracade™ 5222 for herbicide tolerance and insect resistance),• S YN-BTO 11 - 1 x DAS-59122-7 x MIR604 x DAS-01507- 1 x MQN00021 (Agri sure® 3122 for herbicide tolerance and insect resistance),• SYN-BT011-1 x MON00021 (Agrisure™ GT / CB / LL for herbicide tolerance and insect resistance),• SYN-BT011-1 x MIR162 (Agrisure® Viptera™ 2100 for herbicide tolerance and insect resistance),• SYN-BT011-1 x MIR162 x MDN00021 (Agrisure® Viptera™ 3110 for herbicide tolerance and insect resistance),• SYN-BT011-1 x MIR162 x MIR604 (Agrisure® Viptera™ 3100 for herbicide tolerance and insect resistance),• SYN-BT011-1 x MIR162 x DAS-01507-1 x MGN00021 (Agrisure® Viptera™ 3220 for herbicide tolerance and insect resistance),• SYN-BT011-1 x MIR604 (Agrisure™ CB / LL / RW for herbicide tolerance and insect resistance),• SYN-BT011-1 x MIR604 x MDN00021 (Agrisure™ 3000GT for herbicide tolerance and insect resistance), and / or• MIR604 x MON00021 (Agrisure™ GT / RW for herbicide tolerance and insect resistance).DEPOSIT INFORMATION

[0203] A deposit of a representative sample of corn seed containing event MON95275 was made on August 21, 2019, according to the Budapest Treaty with the American Type Culture Collection (ATCC) having an address at 10801 University Boulevard, Manassas, Virginia USA, Zip Code 20110. The deposit was accepted and assigned ATCC Accession No. PTA-126049.EXAMPLES

[0204] The following Examples are included to more fully describe the invention, resulting from the construction and testing of 163 constructs, the production of about 2,300 events, and the analysis of hundreds of thousands of individual plants over 6 years through the rigorous molecular, agronomic, and field testing required for the creation and selection of com event MON95275.

[0205] Those of skill in the art should, in light of the present disclosure, appreciate that many changes can be made in the specific embodiments that are disclosed and still obtain a like or similar result without departing from the spirit and scope of the invention.EXAMPLE 1Expression Cassette Testing, Construct Design, Plant Testing and Construct Selection

[0206] It is often necessary to create and screen a large number of gene expression constructs and transformation events in order to identify a construct, and then an event, which demonstrates optimal expression of the introduced genes of interest, while also not producing agronomic or phenotypic off-types.

[0207] For these reasons, the development of a transgenic corn plant producing insecticidal proteins active against Coleopteran insects without any negative effects on agronomics, yield, or stacking viability, required extensive research, development, and analysis. Specifically, over a 6 year period, over 4,531 proof of concept and commercial transgenic events derived from 163 different plasmid vector constructs were developed, tested, and analyzed.

[0208] This Example describes the design and testing in corn plants of 163 different constructs, to identify the preferred construct for event creation. Each construct varied with respect to the coding sequences for the insecticidal proteins and the transcriptional regulatory elements, and these were tested to select the preferred construct for use in expressing the insecticidal proteins in plants. Each construct had a unique configuration, varying by expression cassette composition (both insecticidal proteins, dsRNAs, and expression elements), orientation, and whether or not proteins were targeted for insertion into chloroplasts.

[0209] In an initial proof of concept and developmental stage, 160 constructs comprising different combinations of 26 distinct promoters, 14 distinct enhancers, 14 distinct introns, 16 distinct insect toxin coding sequences, 16 distinct dsRNA encoding sequences, and 14) distinct 3' UTRs wereused to generate over 2,000 transformed events. After initial molecular characterization for the presence of the transgene(s), 1,875 single transformed com events were selected for further characterization and efficacy testing. These events were evaluated for phenotypic or agronomic off-types, the level of expression of the insect toxin proteins, and efficacy against selected Coleopteran insect pest species. The resulting efficacy and protein expression data, along with any information regarding phenotypic and agronomic off-types, was used to eliminate inefficacious proteins, expression elements, and combinations, and was used to design a smaller number of binary commercial transformation plasmid constructs to be used in the next phase of development. This proof of concept testing stage in the development of MON95275 is identified as “POC Transformation and Assay” in the timeline presented in Figure 3.

[0210] In the next phase of development, 3 new constructs were created. These constructs comprised combinations of 1 to 2 insect toxin transgene expression cassettes and 1 dsRNA expression cassette in different orientations (convergent or divergent). These 3 constructs were used to generate transformed events (also referred to as “transformants”). After shoot formation in culture, a subset of the transformed events were selected based upon visual characteristics and early molecular analysis. After initial transformation, 2,531 transformants were transferred to soil. 1,496 events were discarded after initial molecular characterization. Of the remaining 1,035 events 427 were eliminated based upon observations of plant health. The remaining 608 events were transplanted to pots and grown in the greenhouse (GH) for further assay. Leaf samples of each event were used to measure expression of DvSnf7 specific dsRNA using a QuantiGene ® assay and expressed as femtograms DvSnf7 RNA per total micrograms RNA (fg DvSnf7 / ug RNA). A range of expression of 1,000-3,000 fg DvSnf7 dsRNA / ug RNA was used to select events for further study. Of the remaining 608 events, 425 events were found to express DvSnf7 dsRNA with the range of expression of 1,000-3,000 fg DvSnf7 dsRNA / ug RNA. 19 of the remaining 425 events were eliminated based upon lack of expression of the insect toxin proteins, resulting in a total of 406 events for further assay and characterization. The Ro events were allowed to selfpollinate and produce Ri seed. Based upon observations of plant health and seed return, 152 events were discarded, leaving a total of 254 events for further study. After further molecular characterization, 102 events were discarded, leaving a total of 152 events for Ri nursery for efficacy studies, additional molecular characterization, expression studies, and seed return andsegregation analysis. This commercial transformation and Ro screen stage in the development of event MON95275 is identified as “Comm. TFN Ro Screen” in the timeline presented in Figure 3.

[0211] Since corn rootworm plant assays are destructive assays, requiring the plant to be removed from the pot in order to assess overall root damage, the Ro plants are used to produce Ri seed so that there are sufficient seed to continue generations of each selected event for further efficacy and agronomic assessments, as well as further molecular characterization. Plants derived from the Ri seed were assayed for efficacy against Western Corn Rootworm (Diabrotica virgifera virgifera, WCR) and Northern Corn Rootworm (Diabrotica barberi, NCR). Based upon the efficacy studies, additional molecular characterization, expression studies, and seed return and segregation analysis, 57 events derived from the 3 constructs were advanced for further analysis. This Ri stage efficacy / molecular screen is identified as “GH / Mol. Screen” in Figure 3. After the Ri stage efficacy / molecular screen, events derived from one construct (Construct-2 in Table 2) were discarded based upon decisions regarding the construct configuration and toxin expression cassettes.

[0212] Table 2 shows the number of events remaining corresponding to each construct for each step of selection described above corresponding to each construct. Plasmid construct pM95275 was the construct used in transformation that produced com event MON95275.Table 2. Events per construct selected for continued study.

[0213] The 2017 US field efficacy trials reduced the collective number of events to 14 from Construct-1 and Construct pM95275 based upon efficacy, phenotypic observations, and molecular studies such as insertion site integration.

[0214] Events derived from Construct-1 in Table 2 were discarded based upon decisions regarding insecticidal protein expression.

[0215] Thus, numerous rounds of testing and comparison of various constructs revealed that events produced using the transgene cassette provided as SEQ ID NO: 13, Construct pM95275, provided preferred efficacy against the Coleopteran pest species Western Corn Rootworm (Diabrotica virgifera virgifera, WCR) and Northern Corn Rootworm (Diabrotica barberi, NCR), and preferred molecular characterization and agronomic performance.EXAMPLE 2Field Trials, Molecular Testing and Event Selection

[0216] This Example describes the molecular characterization, analysis, and testing in field trials of events created with Construct pM95275 in multiple locations over several years, which lead to the selection of the final event, MON95275.

[0217] Table 3 illustrates the process used to select the event MON95275. At the commercial transformation Ro screen, one hundred forty (140) Ro transformed events from Construct pM95275 were derived and selected for growth. After quantification of the DvSnf7 expression, 3 events were discarded which did not meet the criteria for an expression range of 1,000-3,000 fg DvSnf7 dsRNA / ug RNA, leaving a total of 137 events for further assay. The remaining 137 events were assayed for expression of the Cry75Aal and Vip4Da2 toxins, and 7 events were discarded based upon the assays, leaving a total of 130 for the Ro screen. After the Ro screen, 52 events were discarded due to poor seed return or plant health, leaving a total of 78 for further molecular characterization. After molecular characterization there were 56 remaining events.

[0218] The remaining 56 events were sent to the Ri nursery for further testing. From the Ri nursery, an additional 8 events were discarded due to poor seed return and segregation analysis and 21 events were discarded due to concerns regarding protein expression and molecular characterization, leaving 19 events.

[0219] The remaining 19 events advanced to the R2 / F1 Cre crossing phase in which the CP4 marker cassette was removed through breeding. During the R2 / F1 Cre crossing phase, 12 events were discarded, 7 due to additional molecular characterization and 5 based upon concerns regarding agronomic performance and other molecular studies.

[0220] The remaining 7 events proceeded to the 2017 U.S. field trials. After the field trials, 3 of the remaining 7 events were discarded, 1 as a result of an off phenotype observed in the field and 2 which performed less well than the others with respect to efficacy and agronomics, leaving 4 events for the 2018 U.S. field trials. During the 2018 US field trials, 1 event was discarded due to an incorrect transcription pattern from the DvSnf7 dsRNA expression cassette and 1 due to agronomic performance, leaving 2 events, Event 1 and event MON95275. After further analysis of the agronomics of the events from multiple field trials in the U.S. and Argentina, event MON95275 was selected as the event for commercialization because it ranked dsRNA higher than Event 1 when all the characteristics of molecular characterization, protein and DvSnf7 dsRNA expression, efficacy and agronomics of each event were compared.Table 3. MON95275 event selection.EXAMPLE 3Cre-excision of the Glyphosate Selection Cassette in Corn Event MON95275

[0221] This Example describes the removal of the glyphosate selection cassette from corn event MON95275 through in vivo Cre-excision. The glyphosate selection cassette was used to select transformed events. By removal of the selection cassette, a “marker-free” event was created wherein only the insecticidal protein expression cassettes remained in the final event.

[0222] Corn variety LH244 immature embryos were transformed using an Agrobacterium- mediated transformation process with Construct pM95275 (presented as SEQ ID NO: 13 and illustrated in Figure 2). Construct pM95275 contains 4 expression cassettes: 2 expression cassettes for the expression of the insecticidal proteins Cry75Aal and Vip4Da2, 1 expression cassette for the expression of the DvSnf7 dsRNA, and a single cassette used for the selection of transformed plant cells using glyphosate selection. The selection cassette was flanked on both sides with Cre- recombinase LoxP recognition sequences.

[0223] After transformation, the Ro transformants were self-pollinated for 2 generations, during which time many events were removed based upon various assays such as efficacy, DvSnf7 expression, protein expression, seed return and plant health, and molecular characterization. By the R2 generation, 19 events remained from the initial 140 events. The 19 homozygous R2 generation events were bred with an elite line of transformed corn plants expressing Cre- recombinase enzyme, derived from Enterobacteria phage Pl.

[0224] This stage in which R2 generation events were bred with plants expressing Cre- recombinase is identified as “Cre Cross” in the timeline presented in Figure 3. Specifically, in this stage, de-tasseled (female) R2 generation plants homozygous for SEQ ID NO: 13 were crosspollinated with transgenic corn plants (male) homozygous for a transgene cassette used for expression of Cre-recombinase enzyme. The Cre-recombinase expressing male donor pollen germinates after landing on the silk tissue of the female plant comprising SEQ ID NO: 13. Once the pollen tube enters the embryo sac, the pollen tube ruptures, setting free the two sperms of the Cre-recombinase expressing male donor. The nucleus of one sperm fuses with the egg nucleus, forming the zygote. The other sperm nucleus fuses with one of the two polar nuclei which in turn fuses with the other polar nucleus, thereby establishing the primary endosperm nucleus.

[0225] Thus, in using the Cre-recombinase expressing plant as the male pollen donor, both the embryo and endosperm of the resulting cross will express Cre-recombinase as the cells divide and develop and become a corn kernel (i.e., seed). The Cre-recombinase binds to inverted repeats in the LoxP site and catalyzes a crossover in an eight-base pair spacer region of the two LoxP sites that flank the expression cassette, resulting in the excision of the marker cassette with one LoxP site remaining in the integrated T-DNA due to recombination (see Figure 2, “Inserted T-DNA After Cre-Excision”).

[0226] The Fi progeny resulting from the Cre Cross were selected for the absence of the CP4 selection cassette and allowed to self-pollinate. This stage in which Fi progeny were allowed to self-pollinate is identified as “Fi Self’ in the timeline presented in Figure 3. Through this process, the two alleles - the Cre-recombinase allele and the allele for the T-DNA used to generate event MON95275 - segregate in the resulting F2 population, resulting in progeny homozygous or heterozygous for one or both alleles.

[0227] The F2 progeny which demonstrated the absence of the Cre-recombinase allele and homozygosity for SEQ ID NO: 9, the transgenic inserted T-DNA after Cre-excision, were selected. These selected F2 progeny were self-pollinated, giving rise to an F3 generation homozygous for SEQ ID NO: 9. This stage in which F2 progeny were allowed to self-pollinate is identified as “F2 Self’ in the timeline presented in Figure 3.

[0228] A further self-pollination resulted in F3 progeny seed (F4 seed) which were assayed for purity and were designated as “Gold Standard Seed.” F4 was the first generation of gold standard seed. Gold Standard Seed is seed that has been assayed for purity to assure the absence of eventsother than MON95275. This stage in which F3 progeny were allowed to self-pollinate is identified as “F3 Self’ in the timeline presented in Figure 3.

[0229] Excision of the glyphosate selection marker cassette did not affect the expression of Cry75Aal, Vip4Da2, and DvSnf7 dsRNA. Removing the glyphosate selection cassette from corn event MON95275 through Cre-excision provided a transgenic corn event which is resistant to Coleopteran pests without adding tolerance to glyphosate in the final event. This “marker-free” event assures flexibility when building com breeding stacks with other com transgenic events to provide a multiplicity of products incorporating event MON95275 and allowing multiple options for providing additional traits in the final commercial breeding stacks.EXAMPLE 4Corn Event MON95275 Demonstrates Resistance to the Coleopteran Insect Pests Western Corn Rootworm and Northern Corn Rootworm

[0230] This Example describes the activity of the com event MON95275 against Coleopteran insect pests. The insect toxin proteins Cry75Aal and Vip4Da2 and the DvSnf7 dsRNA, when expressed together in corn event MON95275, provide resistant to Western Com Rootworm (Diabrotica virgifera virgifera, WCR) and Northern Com Rootworm (Diabrotica barberi, NCR).MON95275 demonstrates resistance to Western Corn Rootworm in the greenhouse and in the field.

[0231] After transformation and insertion of Construct pM95275, Ro stage events were transferred to the greenhouse and allowed to self-pollinate and produce seed. Selected Ri seed was planted in pots and grown in the greenhouse. Eggs from Western Corn Rootworm (WCR) were incubated for approximately 10 days to allow hatching within 4 days after inoculation. 6 plants for each event were assayed. The plants were inoculated at approximately V2 to V3 stage. Each pot was inoculated with about 2,000 eggs. The plants were grown after infestation for approximately 28 days. The plants were then removed from the pots and the roots were carefully washed to remove all soil. The damage to the roots of each plant were assessed using a damage rating scale of 0-3, as presented in Table 4. Comparison was made to a negative wild-type control of the same variety as the transformants. A root damage rating (RDR) of 0-0.75 represents good efficacy, an RDR of 0.76-1.5 represents medium efficacy, and an RDR of 1.6 - 3.0 represents low or poor efficacy.Table 4. Ri root damage rating scores.

[0232] As can be seen in Table 5, com event MON95275 demonstrated significant efficacy when compared to the negative control.Table 5. Average Ri Root Damage (RDR) for corn event MON95275.

[0233] Field efficacy trials were conducted in the United States to assess corn event MON95275 resistance against WCR. Field trials were conducted at 8 separate locations know to have WCR infestations; Colesburg, IA, Fairbank, IA, Independence, IA, Leigh, NE, Pilger, NE, Roanoke, IL, Rowan, IA, and Shelby, NE. Hybrid plants produced by crossing inbred corn event MON95275 (LH244) with corn variety 93 ID 13 were grown in the WCR infested fields. Corn event MON95275 still comprised the CP4 marker cassette in this field trial. In addition, two negative controls were also grown; (1) corn hybrid MON89034 (93IDI3) x LH244 which is Lepidopteran resistant, and (2) non-transgenic corn hybrid 93IDI3 x LH244.

[0234] The trials in each location were planted as a randomized complete block design. The plots were blocked by rep and within that block, plot location was randomized. Both MON95275 and controls were represented once within each block. Block dimensions such as number of columns per row by number of ranges deep varied by location, depending on the size and shape of the field. Each entry was evenly distributed across the field to compensate for any differences in WCRpressure that might occur. Approximately 10 plants each for MON95275 and the controls were dug up at around VT stage. The roots were carefully washed and a Root Damage Ratings (RDR) from 0.1-3.00 was assigned to each plant and is presented in Table 6.Table 6. Field Root Damage Rating (RDR) scale.I l l*For example, if two nodes show 20% and 30% root pnining. the root would be scored as having a root damage rating of 0.50. or if one node shows 10% and two other nodes show 10% each, the root would be scored as having a root damage rating of 0.30. or if two nodes are each missing 50% of their roots, the root would be scored as having a root damage rating of 1.00, etc.

[0235] Tables 7 and 8 show the average Root Damage Ratings for corn event MON95275 and the two negative controls corresponding to each field location.Table 7. Average WCR Root Damage Ratings for MON95275 and controls from Colesburg, IA, Fairbank, IA, Independence, IA, and Leigh, NETable 8. Average WCR Root Damage Ratings for MON95275 and controls from Pilger, NE, Roanoke, IL, Rowan, IA, and Shelby, NE

[0236] As can be seen in Tables 7 and 8, com event MON95275 provided resistance to WCR when compared to the negative controls. While in most cases the controls experienced damage that could potentially lead to economic losses based upon the RDR scale presented in Table 6, corn event MON95275 demonstrated resistance to WCR and only experienced damage that would be considered non-economic across all locations.

[0237] In the summer of 2018 field efficacy trials were conducted in 5 U.S. locations known to have WCR infestations to assess com event MON95275 resistance to WCR; Dundee, IA, Leigh, NE, Oneida, IA, Pilger, NE, and Kingsley, IA. Field trials were conducted and Root Damage Ratings were performed as described above. Both marker-positive and marker-free corn event MON95275 were assayed along with the two negative controls previously described. Tables 9 and 10 show the average Root Damage Ratings for marker and marker-free corn event MON95275 and the two negative controls corresponding to each field location.Table 9. Average WCR Root Damage Ratings for MON95275 and controls from Dundee, IA, Leigh, NE, and Oneida, IA.Table 10. Average WCR Root Damage Ratings for MON95275 and controls from Pilger, NE and Kingsley, IA.

[0238] As can be seen in Tables 9 and 10, both marker-positive and marker-free com event MON 95275 demonstrated resistance to WCR relative to the negative controls. In all but one location, damage to the marker and marker-free corn event MON95275 was non-economic. Damage in Pilger, NE was higher, but still much lower than damage to the negative controls at that location.

[0239] Corn event MON95275 provides resistance to Wester Com Rootworm (Diabrotica virgifera virgifera) as demonstrated in the greenhouse and two U.S. field trials.MON95275 provides resistance to Northern Corn Rootworm in the field

[0240] In the summer of 2017, a single field trial was conducted in Hawkeye, IA, in a field known to be infested with Northern Corn Rootworm (NCR). Marker-positive hybrid corn event MON95275 and the two negative controls as described above were grown on multiple plots in the field in a similar manner as that performed for Western Corn Rootworm. Root Damage Rating were assessed for event MON95275 and the two negative controls using the same scale as that presented in Table 6. Table 11 shows the Average Root Damage Ratings and RDR ranges for the marker-positive corn event MON95275 and the negative controls.Table 11. Average NCR Root Damage Ratings for MON95275 and controls from Hawkeye, IA.

[0241] As can be seen in Table 11 above, the average RDR for marker-positive MON95275 was lower than the MON89034 (93IDI3) x LH244 negative control. The average RDR was low for the non-transgenic control, suggesting NCR pressure was low in the field.

[0242] In the summer of 2018, field trials were conducted in three (3) separate locations known to be infested with NCR, Belmond, IA, Benson, MN, and Colton, SD. Field trials were conducted as previously described. Tables 12 and 13 show the average RDR and RDR range for the three (3) locations.Table 12. Average NCR Root Damage Ratings for MON95275 and controls from Belmond,IA and Benson, MN.Table 13. Average NCR Root Damage Ratings for MON95275 and controls from Colton, SD.

[0243] As can be seen in Tables 12 and 13, corn event MON95275 provided resistance to NCR. For example, in Benson, MN, NCR pressure was high as can be seen in the high average RDRs of the negative controls, but the average RDR was below economic damage in corn event MON95275. In all three (3) locations, MON95275 demonstrated resistance to NCR relative to the two negative controls.

[0244] MON95275 provides resistance to Northern Corn Rootworm (Diabrotica barberi).EXAMPLE 5Corn Event MON95275 Provides Consistent Yield and Similar Agronomics to Untransformed LH244 Corn Plants

[0245] This Example demonstrates that transgenic corn event MON95275 provides consistent yields and similar agronomics in the field to untransformed LH244 com plants.

[0246] Field trials were conducted with plants corresponding to MON95275 prior to Cre-excision of the glyphosate selection cassette and after Cre-excision to determine various aspects of yield and agronomics in comparison to control plants. Measurements of yield were calculated and expressed as bushels per acre (bu / acre). Plant height and ear height were measured in inches (in). 50% pollen shed and 50% silking were expressed as days after planting (DAP).

[0247] In the growing season of 2017 in the United States, yield and agronomic measures were determined for MON95275 inbreds and hybrids pre-Cre-excision of the glyphosate maker cassette. Tables 14 and 15 show the yield and agronomic characteristics measured for MON95275 inbreds and hybrids, respectively. The negative control plants for the inbred comparisons was untransformed variety LH244. Hybrids containing MON95275 were created by cross pollinating the inbred MON95275 with corn variety 93IDI3, and the control was a MON 89034 (93IDI3) x LH244 cross.Table 14. Yield and agronomics for MON95275 inbreds relative to non-transgenic controls.Table 15. Yield and agronomic for MON95275 hybrids relative to non-transgenic controls.

[0248] As can be seen in Tables 15 and 16, the yield and other agronomic measures for MON95275 in the 2017 United States field trials were relatively the same for both inbreds and hybrids relative to the controls. The variability between the inbreds and hybrids and their respective controls was within acceptable limits and demonstrate there were no negative impacts on yield and other agronomic characteristics caused by insertion of the T-DNA into the corn genome of event MON95275.

[0249] In the growing season of 2018 in the United States, yield and agronomic measures were determined for MON95275 inbreds and hybrids pre-Cre-excision and post-Cre-excision of the glyphosate maker cassette. Tables 16 and 17 show the yield and agronomic characteristics measured for MON95275 marker-positive and marker-free inbreds and hybrids, respectively.Table 16. Yield and agronomics for MON95275 inbreds relative to non-transgenic controls.Table 17. Yield and agronomics for MON95275 hybrids relative to non-transgenic controls.

[0250] As can be seen in Tables 16 and 17, the yield and other agronomic measures for MON95275, both marker-positive and marker-free, in the 2018 United States field trials were relatively the same for both inbreds and hybrids relative to the controls. The variability between the inbreds and hybrids and their respective controls was within acceptable limits and demonstrate there were no negative impacts on yield and other agronomic characteristics caused by insertion of the T-DNA into the corn genome of event MON95275.

[0251] Yield and agronomics were also studied in Argentina during the 2018 to 2019 growing season for MON95275 marker-free inbreds. Table 18 shows the yield and agronomic characteristics measured for MON95275 marker-free inbreds.Table 18. Yield and agronomics for MON95275 inbreds relative to non-transgenic controls.

[0252] As can be seen in Table 18, the yield and other agronomic measures were relatively the same for marker-free inbreds and controls from the 2018-2019 Argentina field trials. The variability between the inbreds and the control was within acceptable limits and demonstrate there were no negative impacts on yield and other agronomic characteristics caused by insertion of the T-DNA into the corn genome of event MON95275.EXAMPLE 6Corn Event MON95275 Event-Specific Endpoint TAQMAN® assays

[0253] The following Example describes methods useful in identifying the presence of MON95275 in a corn sample. A pair of PCR primers and a probe were designed for the purpose of identifying the unique junction formed between the corn genomic DNA and the inserted DNA of MON95275 in an event- specific endpoint TAQMAN® PCR. Examples of conditions utilized for identifying the presence of MON95275 in a corn sample in an event-specific endpoint TAQMAN® PCR are described in Table...

Claims

What is claimed is:1 . A recombinant DNA molecule comprising: a) a first nucleotide sequence (i) comprising an expression cassette that encodes a Cry75Aal or Vip4Da2 protein or a DvSnf7 specific dsRNA, and / or (ii) selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7 and SEQ ID NO: 8; and b) a second nucleotide sequence that i. comprises at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 1 1 or 47, or ii. is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8% or at least 99.9% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 11 or 47, or iii. is selected from the group consisting of SEQ ID NOs: 49-148.

2. The recombinant DNA molecule of claim 1, further comprising: c) a third nucleotide sequence that i. comprises at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 12 or 48, or ii. is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8% or at least 99.9% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 12 or 48, or iii. is selected from the group consisting of SEQ ID NOs: 149-248.

3. The recombinant DNA molecule of claim 2, wherein the third nucleotide sequence is selected from the group consisting of SEQ ID NOs: 149-248.

4. The recombinant DNA molecule of any one of claims 1-3, wherein the recombinant DNA molecule has a deletion of one or more consecutive nucleotides between the second nucleotide sequence and the nucleotide sequence relative to SEQ ID NO: 10, 12, or 48.

5. The recombinant DNA molecule of any one of claims 1-4, wherein the second nucleotide sequence is selected from the group consisting of SEQ ID NOs: 49-148.

6. The recombinant DNA molecule of any one of claims 1-5, wherein the recombinant DNA molecule has a deletion of one or more consecutive nucleotides between the first nucleotide sequence and the second nucleotide sequence relative to SEQ ID NO: 10, 11, or 47.

7. A recombinant DNA molecule comprising: a) a first nucleotide sequence (i) comprising an expression cassette that encodes a Cry75Aal or Vip4Da2 protein or a DvSnf7 specific dsRNA, and / or (ii) selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, and SEQ ID NO: 9; and b) a second nucleotide sequence that i. comprises at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 12 or 49, or ii. is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8% or at least 99.9% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 12 or 48, or iii. is selected from the group consisting of SEQ ID NOs: 149-248.

8. The recombinant DNA molecule of claim 7, wherein the second nucleotide sequence is selected from the group consisting of SEQ ID NOs: 149-248.

9. The recombinant DNA molecule of claim 7 or 8, wherein the recombinant DNA molecule has a deletion of one or more consecutive nucleotides between the first nucleotide sequence and the second nucleotide sequence relative to SEQ ID NO: 10, 12, or 48.

10. The recombinant DNA molecule of any one of claims 1-9, further comprising nucleotides 1-1,073 or 15,756-16,861 of SEQ ID NO: 10.

11. The recombinant DNA molecule of any one of claims 1-10, wherein said recombinant DNA molecule is comprised in a corn plant, corn plant part, corn plant cell, corn plant seed, corn progeny plant, or commodity or fuel product made from com and com plant parts.

12. The recombinant DNA molecule of any one of claims 1-10, wherein said recombinant DNA molecule comprises an amplicon diagnostic for the presence of DNA comprising a sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, and SEQ ID NO: 9.

13. The recombinant DNA molecule comprising a polynucleotide of claims 1-11, wherein the recombinant DNA molecule is derived from a corn plant, com plant part, corn seed, processed corn seed, com plant cell or tissue, animal feed comprising com, corn oil, com meal, corn flour, corn flakes, corn bran, food made comprising corn, corn biomass, or fuel products made from corn and corn plant parts.

14. A recombinant DNA molecule comprising a polynucleotide segment of sufficient length to function as a DNA probe that hybridizes specifically under stringent hybridization conditions with a polynucleotide having a sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 47, and SEQ ID NO: 48.

15. A pair of DNA molecules that can function as DNA primers when used together in an amplification reaction comprising a first DNA molecule and a second DNA molecule, wherein the first DNA molecule and the second DNA molecule are different, a) wherein the first DNA molecule is: i. at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, or at least 99.9% identical or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 1, SEQ IDNO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, or SEQ ID NO: 9, or a complement thereof, or ii. at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, or at least 99.9% identical or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 11 or 47, or a complement thereof; and b) wherein the second DNA molecule is: i. at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, or at least 99.9% identical or 100% identical to at least 10, at least 1 1, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, or SEQ ID NO: 9, or a complement thereof, or ii. at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, or at least 99.9% identical or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 11 or 47, or a complement thereof, oriii. at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, or at least 99.9% identical or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 12 or 48, or a complement thereof.

16. The pair of DNA molecules of claim 15, wherein the first DNA molecule and the second DNA molecule can be used together in an amplification reaction with a sample from a plant, plant part, plant seed, plant cell, food or animal feed, or commodity or fuel product made from a plant or plant part to produce an amplicon diagnostic for the presence of a modified corn event MON95275 DNA in said sample.

17. The pair of DNA molecules of claim 16, wherein the amplicon comprises a nucleotide sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10.

18. The pair of DNA molecules of claim 16 or 17, wherein the modified corn event MON95275 DNA is a further modified corn event MON95275 DNA.

19. A method of detecting the presence of a DNA segment diagnostic for a modified corn event MON95275 DNA in a sample, said method comprising: a) contacting said sample with the DNA molecule of any one of claims 1-14; b) subjecting said sample and said DNA molecule to stringent hybridization conditions; and c) detecting hybridization of said DNA molecule to said DNA segment in said sample, wherein said detection is diagnostic for the presence of said modified corn event MON95275 DNA in said sample.

20. A method of detecting the presence of a DNA segment diagnostic for a modified corn event MON95275 DNA in a sample, said method comprising:a) contacting said sample with the pair of DNA molecules of any one of claims 15- 18; b) performing an amplification reaction sufficient to produce a DNA amplicon; and c) detecting the presence of said DNA amplicon in said reaction, wherein the present of said DNA amplicon is diagnostic for the presence of said modified corn event MON95275 DNA in said sample.

21. A method of detecting the presence of a DNA segment diagnostic for a modified corn event MON95275 DNA in a sample, said method comprising performing a sequencing reaction with the sample, wherein the production in the sequencing reaction of a target nucleotide sequence comprising at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, or SEQ ID NO: 9, or a complement thereof, is diagnostic for the modified com event MON95275 DNA in the sample.

22. The method of claim 19, 20, or 21, wherein the modified com event MON95275 is a further modified corn event MON95275.

23. A modified corn plant, corn plant part, com seed, or corn cell comprising a modified corn event MON95275 or the recombinant DNA molecule of any one of claims 1-14.

24. The modified com plant, corn plant part, com seed, or corn cell of claim 23 comprising: a) the recombinant DNA molecule of any one of claims 1-14; or b) a recombinant DNA molecule or DNA segment comprising a nucleotide sequence selected from the group consisting of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10, or a complement thereof; or c) a recombinant DNA molecule or DNA segment comprising a nucleotide sequence that is at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 11, at least12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, at least 50, at least 60, at least 70, at least 80, at least 90, at least 100, at least 150, at least 200, at least 250, at least 300, at least 400, at least 500, at least 1,000, at least 1,500, or at least 2,000 consecutive nucleotides of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, SEQ ID NO: 9, and SEQ ID NO: 10, SEQ ID NO: 11, SEQ ID NO: 12, SEQ ID NO: 47, or SEQ ID NO: 48, or a complement thereof.

25. The modified corn plant, corn plant part, corn seed, or com cell of claim 23 or 24, wherein the corn plant, com plant part, corn seed, or corn cell is further defined as a progeny plant of any generation of a corn plant comprising a modified corn event MON95275, or a com plant part, corn seed, or com cell derived therefrom.

26. The modified corn plant, corn plant part, corn seed, or corn cell of any of claims 23-25, wherein the com plant, corn plant part, corn seed, or com cell exhibits resistance to a Coleopteran insect pest species and / or comprises an expression cassette that encodes a Cry75Aal or Vip4Da2 protein or a DvSnf7 specific dsRNA.

27. The modified corn plant of claims 23-26, wherein the corn plant exhibits resistance to Western Corn Rootworm (Dicibrotica virgifera virgifera, WCR) and / or Northern Corn Rootworm (Diabrotica barberi, NCR).

28. The modified corn plant, corn plant part, corn seed, or corn cell of any of claims 23-27, wherein the recombinant DNA molecule is chromosome 3 of the genome of the modified corn plant, com plant part, com seed, or corn cell, or the DNA segment is present in chromosome 3 of the modified corn plant, corn plant part, corn seed, or corn cell.

29. The modified corn plant, corn plant part, com seed, or corn cell of any one of claims 23- 28, wherein the modified corn event MON95275 is a further modified com event MON95275.

30. The modified corn plant, corn plant part, corn seed, or com cell of claim 23, wherein the modified corn event MON95275 of the modified corn plant, corn plant part, com seed, or corn cell comprises a genetic modification, mutation or edit, relative to the corn event MON95275, introduced via a targeted genome editing technique.31 . A DNA detection kit comprising: a) the recombinant DNA molecule of claim 14; and b) the pair of DNA molecules of any one of claims 15-17.

32. A method of producing a progeny corn plant comprising a modified corn event MON95275 comprising: a) sexually crossing a first modified com plant that comprises a modified corn event MON95275 with itself or a second com plant; b) collecting one or more seeds produced from said cross; c) growing said see to produce one or more progeny plants; and d) selecting at least a first progeny plant or seed comprising a modified corn event MON95275.

33. The method of claim 32, wherein the modified corn event MON95275 is a further modified corn event MON95275.

34. The method of claims 25 or 26, further comprising: e) collecting seed from said at least first progeny plant comprising a modified com event MON95275.

35. A hybrid modified corn plant or seed comprising a modified corn event MON95275 produced by the method of any one of claims 32-34.

36. The hybrid modified com plan or seed of claim 35, wherein the modified corn event MON95275 is a further modified com event MON95275.

37. A nonliving com plant material comprising a detectable amount of the recombinant DNA molecule of any one of claims 1-14.

38. A microorganism comprising the recombinant DNA molecule of any one of claims 1-14.

39. The microorganism of claim 38, wherein the microorganism is a plant cell.

40. A commodity product comprising the recombinant DNA molecule of any one of claims 1- 14.

41. The commodity product of claim 40, wherein said commodity product is produced from a modified com plant, com plant part, corn seed, or corn tissue or cell comprising a modified corn event MON95275.

42. The commodity product of claims 40 or 41, further selected from the group consisting of whole or processed corn seed, animal feed comprising com, com oil, corn meal, corn flour,corn flakes, corn bran, corn biomass, and fuel products produced using corn and com plant parts.

43. The commodity product of any one of claims 40-42, wherein the modified corn event MON95275 is a further modified com event MON95275.

44. A method of producing a commodity product, said method comprising: a) obtaining a modified corn plant, corn plant part, or com seed comprising a modified corn event MON95275; and b) producing a commodity product from the transgenic corn plant, corn plant part, or corn seed.

45. A corn plant, corn plant part, or corn seed comprising a DNA molecule or segment functional as a template when tested in a DNA amplification method to produce an amplicon diagnostic for the presence of a modified com event MON95275 DNA.

46. A method of determining the zygosity of a corn plant corn plant part, or corn seed comprising a modified corn event MON95275 comprising: a) contacting a sample comprising DNA from the corn plant, com plant part, or corn seed with the pair of DNA molecules of claim 15; b) performing a nucleic acid amplification reaction with the sample and the pair of DNA molecules; and c) detecting in the nucleic acid amplification reaction a first amplicon diagnostic for a modified corn event MON95275 and a second amplicon diagnostic for native corn genomic DNA not comprising the modified corn event MON95275, wherein the presence of only the first amplicon is diagnostic of a com plant, com plant part, or corn seed homozygous for the modified corn event MON95275, and the presence of both the first amplicon and the second amplicon is diagnostic of a corn plant, corn plant part, or com seed heterozygous for the modified corn event MON95275.

47. A method of determining the zygosity of a com plant, corn plant part, or corn seed comprising a modified corn event MON95275 comprising: a) contacting a sample comprising DNA from the corn plant, corn plant part, or corn seed with a first primer pair that can produce a first amplicon of all or part of the modified corn event MON95275 and a second primer pair that can produce asecond amplicon of a standard genomic sequence known to be single copy and homozygous in the corn plant, com plant part, or com seed; b) contacting the sample with a first probe that specifically hybridizes to the first amplicon and / or all or part of the modified corn event MON95275, and a second probe that specifically hybridizes to the standard genomic sequence; c) performing a DNA amplification reaction using real-time PCR with the sample and determining the cycle thresholds (Ct values) of the first amplicon and the second amplicon; d) calculating the difference (ACt) between the Ct values of the second amplicon and the first amplicon; and e) determining the zygosity of the modified corn event MON95275, wherein a ACt of about zero (0) indicates homozygosity of the modified corn event MON95275 and a ACt of about one (1) indicates heterozygosity of the modified corn event MON95275.

48. The method of claim 46 or 47, wherein the first and second primer pairs comprise SEQ ID NO: 15 combined with SEQ ID NO: 16, and SEQ ID NO: 18 combined with SEQ ID NO: 19.

49. A method of determining the zygosity of a com plant, corn plant part, or corn seed comprising a modified corn event MON95275 comprising: a) contacting a sample comprising DNA from the corn plant, com plant part, or corn seed with a primer pair capable of producing a first amplicon diagnostic for the modified corn event MON95275 and a second amplicon diagnostic for native corn genomic DNA not comprising the modified corn event MON95275; b) performing a nucleic acid amplification reaction with the sample and the set of primer pairs; and c) detecting the first amplicon and the second amplicon, wherein the presence of only the first amplicon is diagnostic of a corn plant, com plant part, or com seed homozygous for the modified corn event MON95275, the presence of only the second amplicon is diagnostic of a corn plant, corn plant part, or com seed homozygous for native corn genomic DNA not comprising the modified corn event MON95275, and the presence of both the first amplicon and the second ampliconis diagnostic of a com plant, corn plant part, or com seed heterozygous for the modified corn event MON95275.

50. A method for determining the zygosity of a corn plant, corn plant part, or corn seed comprising a modified corn event MON95275 comprising: a) contacting a sample comprising DNA from the corn plant, corn plant part, or corn seed with a probe set which contains at least a first probe that specifically hybridizes to the modified com event MON95275 and at least a second probe that specifically hybridizes to corn genomic DNA that was disrupted by insertion of the heterologous DNA of corn event MON95275 and is disrupted by the modified corn event MON95275 DNA, wherein the second probe does not hybridize to the modified corn event MON95275 DNA; and b) hybridizing the probe set with the sample under stringent hybridization conditions, wherein detecting hybridization of only the first probe under the hybridization conditions is diagnostic for a com plant, corn plant part, or com seed homozygous for the modified com event MON95275, and wherein detecting hybridization of both the first probe and the second probe under the hybridization conditions is diagnostic for a com plant, corn plant part, or corn seed heterozygous for the modified corn event MON95275.

51. The method of claim 50, wherein the probe set comprises SEQ ID NO: 17 and SEQ ID NO: 20.

52. The method of any one of claims 46-51, wherein the modified corn event MON95275 is a further modified com event MON95275.

53. A population of transgenic com plants, wherein each transgenic corn plant comprises a modified corn event MON95275.

54. The population of claim 53, wherein each transgenic com plant of the population exhibits resistance to a Coleopteran insect pest species.

55. The population of transgenic com plants of claim 53 or 54, wherein the modified com event MON95275 is a further modified com event MON95275.

56. A method of modifying a com plant, the method comprising: a) introducing a site-specific nuclease or a recombinant DNA construct comprising an expression cassette encoding a sit-specific nuclease into at least one cell of anexplant of a corn plant comprising com event MON95275, or a plant part thereof, to produce a modified corn event MON95275 via a genome editing technique; and b) developing or regenerating a modified com plant from the explant, wherein the modified corn plant comprises the modified corn event MON95275.

57. The method of claim 56, wherein the site-specific nuclease is a zinc-finger nuclease (ZFN), a meganuclease, an RNA-guided endonuclease, a TALE-endonuclease (TALEN), a recombinase, or a transposase.

58. The method of claim 56 or 57, wherein the site-specific nuclease is an RNA-guided endonuclease or a CRISPR / Cas nuclease.

59. The method of claim 58, wherein the introducing step (a) comprises introducing the recombinant DNA construct into the at least one cell of the explant, and wherein the recombinant DNA construct further comprises an expression cassette encoding a first guide RNA (gRNA).

60. The method of claim 59, wherein the recombinant DNA construct further comprises an expression cassette encoding a second guide RNA (gRNA).

61. The method of claim 58, wherein the introducing step (a) further comprises introducing a first guide RNA (gRNA) or a second recombinant DNA construct comprising an expression cassette encoding a first guide RNA (gRNA) into the at least one cell of the explant.

62. The method of claim 61, wherein the introducing step (a) comprises introducing at least two guide RNAs (gRNAs) comprising the first gRNA and a second gRNA into the at least one cell of the explant.

63. The method of claim 61, wherein the second recombinant DNA construct further comprises an expression cassette encoding a second guide RNA (gRNA).

64. The method of claim 61, wherein the introducing step (a) comprises introducing a first gRNA and second recombinant DNA construct comprising an expression cassette encoding a second guide RNA (gRNA) into the at least one cell of the explant.

65. The method of any one of claims 56-64, wherein the site-specific nuclease has a first target site in the genome of the corn plant at or near com event MON95275.

66. The method of claim 65, wherein the site-specific nuclease has a second target site in the genome of the com plant at or near com event MON95275.

67. The method of any one of claims 56-66, wherein the introducing step (a) comprises introducing a second site-specific nuclease or a recombinant DNA construct comprising an expression cassette encoding a second site specific nuclease into at least one cell of the explant, and wherein the second site-specific nuclease has a second target site in the genome of the com plant at or near corn event MON95275.

68. The method of any one of claims 59-64, wherein the first gRNA has a first target site in a flanking DNA sequence, 5' flank, 3 ' flank, junction sequence, or insertion sequence of corn event MON95275, or a complement thereof.

69. The method of any one of claims 59-64 and 68, wherein the first gRNA has a first target site comprising a target sequence that is: i. at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 11 or 47, or a complement thereof; or ii. at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 12 or 48, or a complement thereof; or iii. at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 1 1, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least18, at least 19, at least 20, at least 21 , at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, or SEQ ID NO: 9, or a complement thereof.

70. The method of claim 69, wherein the first gRNA has a second target site comprising a target sequence that is: i. at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 11 or 47, or a complement thereof; or ii. at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 1 1, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 12 or 48, or a complement thereof; or iii. at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4,SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, or SEQ ID NO: 9, or a complement thereof.

71. The method of any one of claims 59-64 and 68-70, i. wherein the first gRNA has a first target site comprising a target sequence that is:

1. at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 1 1 or 47, or a complement thereof; or2. at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 12 or 48, or a complement thereof; or3. at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3,SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, or SEQ ID NO: 9, or a complement thereof; and ii. wherein the second gRNA has a second target site comprising a target sequence that is:

1. at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 11 or 47, or a complement thereof; or2. at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 1 1, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 12 or 48, or a complement thereof; or3. at least 90%, at least 91%, at least 92%, at least 93%, at least 94%, at least 95%, at least 96%, at least 97%, at least 98%, at least 99%, at least 99.1%, at least 99.2%, at least 99.3%, at least 99.4%, at least 99.5%, at least 99.6%, at least 99.7%, at least 99.8%, at least 99.9% or 100% identical to at least 10, at least 11, at least 12, at least 13, at least 14, at least 15, at least 16, at least 17, at least 18, at least 19, at least 20, at least 21, at least 22, at least 23, at least 24, at least 25, at least 30, at least 35, at least 40, at least 45, or at least 50 consecutive nucleotides of SEQ ID NO: 1, SEQ ID NO: 2, SEQ ID NO: 3, SEQ ID NO: 4, SEQ ID NO: 5, SEQ ID NO: 6, SEQ ID NO: 7, SEQ ID NO: 8, or SEQ ID NO: 9, or a complement thereof.

72. The method of any one of claims 66, 67, 70, or 71, wherein the modified corn event MON95275 comprises a deletion or excision of intervening genomic DNA between the first target site and the second target site, relative to the corn event MON95275.

73. The method of any one of claims 56-72, further comprising: c) selecting the modified corn plant comprising the modified corn event MON95275; and d) sexually crossing the modified corn plant with itself or a second com plant to produce one or more modified progeny corn plants.

74. A method of introducing a target site into a corn plant, the method comprising: a) introducing a cognate target site into the corn event MON95275 locus of at least one cell of a corn plant or corn plant part comprising the corn event MON95275 or an explant thereof via a targeted genome editing technique, wherein the cognate target site is identical or similar to an originator target site for a site-specific nuclease present in the com event MON95275 locus; and b) developing or regenerating a modified corn plant comprising a modified corn event MON95275 comprising the cognate target site.

75. The method of claim 74, further comprising: c) introducing a site-specific nuclease or a recombinant DNA construct comprising an expression cassette encoding a site specific nuclease into at least one cell of an explant of a corn plant comprising the modified corn event MON95275 or a plant part thereof, to produce a further modified corn event MON95275 via a targeted genome editing technique, wherein the target site of the site-specific nuclease includes the cognate target site and the originator target site; and d) developing or regenerating a second modified corn plant comprising the further modified corn event MON95275.

76. A method of introducing a target site into a corn plant, the method comprising: a) introducing a cognate target site into the corn event MON95275 locus of at least one cell of a corn plant or corn plant part comprising the corn event MON95275 or an explant thereof via a targeted genome editing technique to produce a modified corn event MON95275 comprising the cognate target site, wherein the cognatetarget site is identical or similar to an originator target site for a site-specific nuclease present in the corn event MON95275 locus, and b) introducing a site-specific nuclease or a recombinant DNA construct comprising an expression cassette encoding a site specific nuclease into at least one cell of an explant of a modified corn plant comprising the modified corn event MON95275, or a plant part thereof, to produce a further modified corn event MON95275 via a targeted genome editing technique, wherein the target site of the site-specific nuclease includes the cognate target site and the originator target site; and c) developing or regenerating a second modified com plant comprising the further modified corn event MON95275.

77. The method of claim 75 or 76, wherein the further modified corn event MON95275 of the second modified corn plant comprises a deletion or excision of intervening genomic DNA between the originator target site and the cognate target site, relative to the corn event MON95275 or modified com event MON95275.

78. The method of claim 15-7 , further comprising: selecting the second modified corn plant or a progeny plant of the second modified com plant comprising the further modified corn event MON95275, and sexually crossing the second modified com plant or the progeny plant with itself or another com plant to produce one or more modified progeny corn plants comprising the further modified com event MON95275.

79. The method of claim 74 or 76, wherein the modified corn event MON95275 is a further modified corn event MON95275.

80. A method of producing a progeny corn plant comprising a modified corn event MON95275 comprising: a) sexually crossing a first modified com plant that comprises a modified corn event MON95275 with itself or a second com plant; b) collecting one or more seeds produced from said cross; c) growing said seed to produce one or more progeny plants; and d) selecting at least a first progeny plant or seed comprising a modified corn event MON95275.

81. A method of modifying an explant of a corn plant or plant part, the method comprising: introducing a site-specific nuclease or a recombinant DNA construct comprising an expression cassette encoding a site-specific nuclease into at least one cell of an explant of a com plant or plant part comprising corn event MON95275 to produce a modified corn event MON95275 into the at least one cell of the explant.

82. The method of claim 81, wherein the modified corn event MON95275 is a further modified corn event MON95275.

Images