US 12,467,061 B2
Plant pathogen effector and disease resistance gene identification, compositions, and methods of use
James Cahill, Des Moines, IA (US); April L Leonard, Des Moines, IA (US); Bailin Li, Johnston, IA (US); Yurong Li, Johnston, IA (US); and Shawn Thatcher, Urbandale, IA (US)
Assigned to PIONEER HI-BRED INTERNATIONAL, INC., Johnston, IA (US)
Appl. No. 17/995,707
Filed by PIONEER HI-BRED INTERNATIONAL, INC., Johnston, IA (US)
PCT Filed Mar. 9, 2021, PCT No. PCT/US2021/021505
§ 371(c)(1), (2) Date Oct. 7, 2022,
PCT Pub. No. WO2021/211227, PCT Pub. Date Oct. 21, 2021.
Claims priority of provisional application 63/010,452, filed on Apr. 15, 2020.
Claims priority of provisional application 63/010,462, filed on Apr. 15, 2020.
Prior Publication US 2023/0151382 A1, May 18, 2023
Int. Cl. C12N 15/82 (2006.01); C07K 14/375 (2006.01)
CPC C12N 15/8282 (2013.01) [C07K 14/375 (2013.01)] 2 Claims
 
1. A method of validating the presence of a gene that confers resistance to disease caused by Puccinia polysora, the method comprising:
a. identifying at least one potential gene or disease resistance locus in a maize plant that confers resistance to disease caused by Puccinia polysora;
b. transfecting at least one allele of a plant pathogen effector gene and a luciferase gene into a maize protoplast, wherein the maize protoplast is derived from the disease resistant plant, and wherein the pathogen effector gene encodes a polypeptide comprising an amino acid sequence of at least 95% sequence identity, when compared to any one of SEQ ID NOs: 12, 2, 4, 10, 11, and 13-23;
c. expressing the at least one allele of a plant pathogen effector gene and the luciferase gene before measuring luciferase activity; and
d. validating the presence of a gene in the protoplast that produces a hypersensitive response in the presence of the plant pathogen effector as indicated by reduced luciferase activity in the protoplast relative to a control protoplast which is transfected with the luciferase gene and which is not transfected with the at least one allele of a plant pathogen effector.