Wheat having reduced waxy protein due to non-transgenic alterations of a waxy gene

8,735,649

11/004536

December 03, 2004

A series of independent non-transgenic mutations found at the waxy loci of wheat; wheat plants having these mutations in their waxy loci; and a method of creating and finding similar and/or additional mutations of the waxy by screening pooled and/or individual wheat plants. The wheat plants of the present invention exhibiting altered waxy activity in the wheat without having the inclusion of foreign nucleic acids in their genomes. The invention also includes food and non-food products as well as non-food products that incorporate seeds from the wheat plants having non-transgenic mutations in one or more waxy genes.

Fuerstenberg, Susan I (Hercules, CA, US); Slade, Ann J. (Kenmore, WA, US)

Arcadia Biosciences, Inc. (Davis, CA, US)

1. A method of producing a tetraploid or hexaploid wheat plant, or part thereof, exhibiting reduced waxy enzyme activity compared to a wild type wheat plant or part thereof, said method comprising the steps of: a) obtaining plant material from a parent wheat plant, wherein the parent wheat plant comprises two or more wild-type waxy genes; b) creating mutagenized plant material by inducing at least one mutation in each of the at least two waxy genes of the parent plant material, wherein said mutations are induced by treating the wheat plant material with a mutagen; c) analyzing the mutagenized wheat plant material, or a progeny wheat plant produced from the mutagenized wheat plant material, to identify a plant having at least one mutation in each of the at least two waxy genes, wherein said analysis is performed by isolating genomic DNA from the mutagenized wheat plant material, or a progeny wheat plant produced from the mutagenized wheat plant material, and amplifying segments of a waxy gene in the isolated genomic DNA by using primers specific to the waxy gene or to the DNA sequences adjacent to the waxy gene, wherein one of the at least two mutated waxy genes being identified encodes a protein comprising a A468V mutation in SEQ ID NO: 4, and one of the at least two mutated waxy genes being identified encodes a protein comprising a Q197* truncation mutation in SEQ ID NO: 6; and d) producing a tetraploid or hexaploid progeny wheat plant from the mutagenized wheat plant material or the progeny wheat plant produced from the mutagenized wheat plant material, each comprising the at least two mutated waxy genes as identified in step c), wherein the A468V mutation in SEQ ID NO: 4 and the Q197* truncation mutation in SEQ ID NO: 6 cause a reduction waxy enzyme activity in the progeny wheat plant compared to the parent wheat plant of step a), as indicated by a reduced level of amylose in the progeny wheat plant or a part thereof.

2. The method of claim 1 wherein the wheat plant material is selected from the group consisting of seeds, pollen, plant cells, and plant tissue.

3. The method of claim 1 wherein the mutagen is ethyl methanesulfonate.

4. The method of claim 3 wherein the concentration of ethyl methanesulfonate used is from about 0.75% to about 1.2%.

5. The method of claim 1 wherein analyzing the mutagenized plant material or a progeny wheat plant produced from the mutagenized wheat plant material further comprises analysis of a starch characteristic.

6. The method of claim 5 wherein the analysis of a starch characteristic comprises iodine staining.

7. The method of claim 1 wherein the part thereof is a seed.

800/270

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