# US 12,215,330 B2 · Wheat with reduced lipoxygenase activity

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- Number: US 12,215,330 B2
- Document ID: US12215330B2
- Status: Granted
- Kind: B2
- Application number: 18/145,611
- Filed: 2022-12-22
- Published: 2023-07-06
- Granted: 2025-02-04
- Earliest claimed: 2015-06-19
- Assignees: Arcadia Biosciences, Inc.
- Applicants: Arcadia Biosciences, Inc.
- Inventors: Ann Slade; Michelle Noval; Dayna Loeffler; Jessica Mullenberg; Aaron Holm
- Web page: https://patentgrep.com/patent/US12215330B2

## CPC

- C12N 15/8247: {involving modified lipid metabolism, e.g. seed oil composition}
- A01H 5/10: Seeds
- A01H 6/4678: {Triticum sp. [wheat]}
- A23L 7/10: Cereal-derived products
- A23L 7/198: {Dry unshaped finely divided cereal products, not provided for in groups A23L7/117 - A23L7/196 and A23L29/00, e.g. meal, flour, powder, dried cereal creams or extracts}
- C12N 9/0069: {acting on single donors with incorporation of molecular oxygen, i.e. oxygenases (1.13)}
- C12N 15/8243: {involving biosynthetic or metabolic pathways, i.e. metabolic engineering, e.g. nicotine, caffeine}
- C12Y 113/11: with incorporation of two atoms of oxygen (1.13.11)
- C12Y 113/11012: Linoleate 13S-lipoxygenase (1.13.11.12)
- A23V 2002/00: Food compositions, function of food ingredients or processes for food or foodstuffs
- C12N: MICROORGANISMS OR ENZYMES; COMPOSITIONS THEREOF; PROPAGATING, PRESERVING, OR MAINTAINING MICROORGANISMS; MUTATION OR GENETIC ENGINEERING; CULTURE MEDIA (microbiological testing media C12Q1/00)
- A01H: NEW PLANTS OR {NON-TRANSGENIC} PROCESSES FOR OBTAINING THEM; PLANT REPRODUCTION BY TISSUE CULTURE TECHNIQUES
- A23L: FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES, NOT OTHERWISE PROVIDED FOR; PREPARATION OR TREATMENT THEREOF (preservation thereof A23B)
- C12Y: ENZYMES
- A23V: INDEXING SCHEME RELATING TO FOODS, FOODSTUFFS OR NON-ALCOHOLIC BEVERAGES AND LACTIC OR PROPIONIC ACID BACTERIA USED IN FOODSTUFFS OR FOOD PREPARATION

## Abstract

A series of independent human-induced non-transgenic mutations found at one or more of the Lpx genes of wheat; wheat plants having these mutations in one or more of their Lpx genes; and a method of creating and finding similar and/or additional mutations of Lpx by screening pooled and/or individual wheat plants. The wheat plants disclosed herein exhibit decreased lipoxygenase activity without having the inclusion of foreign nucleic acids in their genomes. Additionally, products produced from the wheat plants disclosed herein display increased oxidative stability and increased shelf life without having the inclusion of foreign nucleic acids in their genomes.

## Claims

### Claim 1

A wheat flour comprising a human induced alteration in an Lpx1 gene in a D genome, wherein the wheat flour has a property selected from the group consisting of: (a) increased shelf-life; (b) increased oxidative stability; (c) decreased production of Lpx1 protein; (d) decreased lipoxygenase activity; (e) decreased hexanal production; (f) decreased pinellic acid production; (g) decreased decomposition products from fatty acids; and (h) improved sensory characteristics as compared to flour from a wild type wheat grain.

### Claim 2 (depends on 1)

The wheat flour of claim 1, wherein the Lpx1 gene is Lpx-D1.

### Claim 3 (depends on 2)

The wheat flour of claim 2, wherein the human-induced alteration in the Lpx-D1 gene is homozygous.

### Claim 4 (depends on 2)

The wheat flour of claim 2, wherein the human induced alteration in the Lpx-D1 gene is in a promoter.

### Claim 5 (depends on 2)

The wheat flour of claim 2, wherein the human induced alteration in the Lpx-D1 gene results in a premature stop codon.

### Claim 6 (depends on 2)

The wheat flour of claim 2, wherein the human induced alteration in the Lpx-D1 gene results in:
(i) a change from a tryptophan to a stop codon at amino acid position 81 (W81*), 101 (W101*), 494 (W494*), 517 (W517*), 591 (W591*), 635 (W635*), 665 (W665*), 666 (W666*), 701 (W701*), or 789 (W789*) of SEQ ID No. 3;
(ii) a change from a glutamine to a stop codon at amino acid position 190 (Q190*) of SEQ ID No. 3; or
(iii) a change from a guanine to adenine at nucleotide position 2386 (G2386A), or 2827 (G2827A) of SEQ ID No. 1; or a change from a guanine to adenine at nucleotide position 1538 (G1538A) of SEQ ID No. 15.

### Claim 7 (depends on 1)

The wheat flour of claim 1, further comprising a human induced alteration in a Lpx1 gene of a B genome.

### Claim 8 (depends on 7)

The wheat flour plant of claim 7, wherein the Lpx1 gene of the B genome is Lpx-B1.2.

### Claim 9 (depends on 8)

The wheat flour of claim 8, wherein the mutation in the B genome of the Lpx-B1.2 gene results in a change from a tryptophan to a stop codon at amino acid position 510 (W510*) of SEQ ID No. 6.

### Claim 10 (depends on 1)

The wheat flour of claim 1, wherein the flour is selected from the group consisting of: a whole grain flour, an ultrafine milled whole grain flour, a coarse fraction, a bran, a refined flour, and combinations thereof.

### Claim 11 (depends on 1)

The wheat flour of claim 1, wherein the flour has an increased shelf-life of at least 1 month as compared to flour from a wild type wheat grain.

### Claim 12 (depends on 1)

The wheat flour of claim 1, wherein the flour has at least a 5% reduction in hexanal production as compared to flour from a wild type wheat grain.

### Claim 13 (depends on 1)

The wheat flour of claim 1, wherein the flour has at least a 9% reduction of lipoxygenase activity as compared to flour from a wild type wheat grain.

### Claim 14 (depends on 11)

The wheat flour of claim 11, wherein the flour has at least a 60% reduction of lipoxygenase activity as compared to flour from a wild type wheat grain.

### Claim 15

A wheat flour made from wheat grain comprising a human induced alteration in an Lpx1 gene in each of a B and a D genome, wherein the flour has a property selected from the group consisting of: (a) increased shelf-life; (b) increased oxidative stability; (c) decreased production of Lpx1 protein; (d) decreased lipoxygenase activity; (e) decreased hexanal production; (f) decreased pinellic acid production; (g) decreased decomposition products from fatty acids; and (h) improved sensory characteristics as compared to flour from a wild type wheat grain.

### Claim 16 (depends on 15)

The wheat flour of claim 15, wherein the Lpx1 gene in the D genome is Lpx-D1.

### Claim 17 (depends on 16)

The wheat flour of claim 16, wherein the human induced alteration in the Lpx-D1 gene results in a premature stop codon.

### Claim 18 (depends on 16)

The wheat flour of claim 16, wherein the human induced alteration in the Lpx-D1 gene results in:
(i) a change from a tryptophan to a stop codon at amino acid position 81 (W81*), 101 (W101*), 494 (W494*), 517 (W517*), 591 (W591*), 635 (W635*), 665 (W665*), 666 (W666*), 701 (W701*), or 789 (W789*) of SEQ ID No. 3;
(ii) a change from a glutamine to a stop codon at amino acid position 190 (Q190*) of SEQ ID No. 3; or
(iii) a change from a guanine to adenine at nucleotide position 2386 (G2386A), or 2827 (G2827A) of SEQ ID No. 1; or a change from a guanine to adenine at nucleotide position 1538 (G1538A) of SEQ ID No. 15.

### Claim 19 (depends on 15)

The wheat flour of claim 15, wherein the Lpx1 gene in the B genome is Lpx-B1.2.

### Claim 20 (depends on 15)

The wheat flour of claim 15, wherein the flour has an increased shelf-life of at least 1 month as compared to flour from a wild type wheat grain.

### Claim 21 (depends on 15)

The wheat flour of claim 15, wherein the flour has at least a 5% reduction in hexanal production as compared to flour from a wild type wheat grain.

### Claim 22 (depends on 15)

The wheat flour of claim 15, wherein the flour has at least a 9% reduction of lipoxygenase activity as compared to flour from a wild type wheat grain.