Methods for generating and identifying mutant polyploid plants, and uses therefor

• US 6,696,294 B1
• Filed: 12/18/2000
• Issued: 02/24/2004
• Est. Priority Date: 06/19/1998
• Status: Expired due to Fees

• Current Assignee: Omega Genetics LLC
• Original Assignee: Northwest Plant Breeding Co. (Corteva, Inc.)
• Inventors: Konzak, Calvin F.
• Primary Examiner(s): Grunberg, Anne Marie
• Application Number:
  US09/719,880
• Time in Patent Office: 1,163 Days
• Field of Search: 800/276, 800/270, 800/260, 800/295, 800/298, 800/320, 800/320.3, 435/410, 435/444, 435/447, 435/446, 435/441, 435/440
• US Class Current: 435/441
• CPC Class Codes: A01H 1/06 Processes for producing mutations, e.g. treatment with chemicals or with radiation specific mutations prepared by genetic engineering on plant cell or plant tissues C12N15/00 ; process for producing transgenic plants C12N15/82

Abstract

The present invention provides methods for generating and identifying mutations in any target gene of a polyploid plant species. In a preferred aspect of the present invention, a plant is constructed and/or selected that has at least one copy of a functional, target gene located exclusively in only one of its homoeologous, or homologous, genomes. Seed derived from the selected plant are then contacted with an effective amount of at least one mutagenic agent, the treated seed are germinated and the seeds or plants derived therefrom, are screened for mutations in the target gene. Thus, the inventive concepts set forth herein can be used to create, select and identify mutations in any target gene of any suitable polyploid plant, thereby providing a source of numerous, readily-identifiable mutations that can, if so desired, be used in crosses to develop unique new crop cultivars. New alleles of homologous or homoeologous target genes can be recombined to generate an almost unlimited range of genetically-controlled novel phenotypes having predetermined, desirable properties.

Claims

1. A method for producing mutants of a target gene in a wheat plant, comprising;

constructing a wheat plant having at least one functional copy of a target gene located exclusively in only one of the homoeologous or homologous genomes of said wheat plant;

consecutively contacting seeds derived from the constructed wheat plant with an effective amount of at least two mutagenic agents to yield mutagenized seeds;

germinating said mutagenized seeds; and

assaying seed from wheat plants derived from said germinated, mutagenized seeds to identify mutants of the target gene.

2. The method of claim 1 wherein the seeds derived from the constructed wheat plant are contacted with a chemical mutagenic agent.

3. The method of claim 2 wherein the chemical mutagenic agent is selected from the group consisting of ethyl methanesulfonate, diethyl sulfate and sodium azide.

4. The method of claim 1 wherein the seeds derived from the constructed wheat plant are contacted with a first chemical mutagenic agent selected from the group consisting of ethyl methanesulfonate and diethyl sulfate, and are then contacted with sodium azide.

5. The method of claim 4 wherein the seeds are soaked in water for from about four to about six hours before being contacted with a chemical mutagenic agent.

6. The method of claim 4 wherein ethyl methanesulfonate is utilized at a concentration of from about 0.25 milliliters to about 0.35 milliliters per liter of water.

7. The method of claim 6 wherein the seeds are contacted with ethyl methanesulfonate for a period of from about two hours to about two and a half hours.

8. The method of claim 4 wherein diethyl sulfate is utilized at a concentration of from about 1.0 milliliters to about 2.0 milliliters per liter of water.

9. The method of claim 8 wherein the seeds are contacted with diethyl sulfate for a period of from about two hours to about two and a half hours.

10. The method of claim 4 wherein sodium azide is utilized at a concentration of from about 1×

10<sup>−

3</sup>M to about 2'"'"'10<sup>−

3</sup>M.

11. The method of claim 10 wherein the seeds are contacted with sodium azide for a period of from about one hour to about one and a half hours.

12. The method of claim 1 wherein the mutagenized seeds are germinated within about two hours after being contacted with an effective amount of at least two mutagenic agents.

13. The method of claim 1 wherein said target gene is involved in starch biosynthesis.

14. The method of claim 1 wherein said target gene is involved in phytic acid biochemistry.

15. The method of claim 1 wherein said target gene encodes a lipase.

16. The method of claim 1 wherein said target gene encodes a lipoxygenase.

17. The method of claim 1 wherein said target gene encodes a polyphenol oxygenase.

18. A method for producing mutants of a target gene in a wheat plant, comprising;

constructing a wheat plant having at least one functional copy of a target gene located exclusively in only one of the homoeologous or homologous genomes of said wheat plant;

consecutively contacting seeds derived from the constructed wheat plant with an effective amount of at least two mutagenic agents to yield mutagenized seeds;

germinating said mutagenized seeds; and

assaying wheat plants derived from said germinated, mutagenized seeds to identify mutants of the target gene.