Project Details:

Title:
Characterization and enhancement of soybean genetic resources for soilborne disease resistance

Parent Project: Characterization and enhancement of soybean genetic resources for soilborne disease resistance
Checkoff Organization:North Central Soybean Research Program
Categories:Breeding & genetics, Soybean diseases
Organization Project Code:NCSRP
Project Year:2017
Lead Principal Investigator:James Kurle (University of Minnesota)
Co-Principal Investigators:
Jianxin Ma (Purdue University)
Aaron Lorenz (University of Minnesota)
Nevin Young (University of Minnesota)
Keywords: Genetic soilborn disease

Contributing Organizations

Funding Institutions

Information and Results

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Project Summary

The goals of this project are to characterize, identify, isolate , and utilize novel genes/QTLs conferring complete resistance or partial resistance to P. sojae, P. ultimum P. irregulare, and F. graminearum. The new sources of resistance will then be deployed for effective disease management in the North Central region in collaboration with the USB-funded soybean disease research project. We have four research objectives that will help us achieve these goals

Project Objectives

Project Deliverables

(1) Markers/QTLs for partial resistance to root rot caused by P. sojae, F. graminearum, P. ultimum, and P. irregulare. Markers/ QTLs associated with resistance to seed rot and damping off caused by P. ultimum and P. sojae. Gene-based markers specific to RpsUN1 and RpsUN2.
(2) Early maturity lines or pre-breeding experimental lines resistant to P. sojae, F. graminearum, P. ultimum, and P. irregulare that can be used in breeding commercial soybean cultivars adapted to northern soybean growing areas.

Progress of Work

Update:
Objective 1:
Seed, seedling, and root rot caused by F. graminearum and P. ultimum var.ultimum was evaluated in six experiments conducted to 1) confirm repeatability of results obtained in inoculated assays (three experiments) and 2) to phenotype all 280 ancestral lines for resistance to F. gramearum, P.ultimum, and P. irregulare seedling rot (three experiments). In preparation for phenotyping the ancestral lines, the response of a selected subset consisting of 24 lines of the 280 ancestral lines were evaluated for their response to the seed and seedling rot caused by inoculation with Pythium ultimum and by inoculation with P. irregulare. The evaluations were conducted twice with three reps for both pathogens. The seed rot assay results were compared by correlation analysis to determine the repeatability of the results obtained using the in vitro inoculation method. Correlation analysis of results within each Pythium species indicated that root length or root mass yielded the most consistent responses to inoculation (P. ultimum : r2 of 0.87 and 0.91 for root length and root mass respectively). Response to seed root (in vitro assay) was not correlated with the response to seedling rot. Three replicates of 280 lines are being evaluated for resistance response to inoculation with P. ultimum and to inoculation with P irregulare in the seedling assay planted in growth media. This trial was just completed and inoculated plants are being evaluated and assigned a root necrosis score and the ratio of uninoculated to inoculated root length and root dry weight are being calculated and data recorded. Three replicates of the seed assay for resistance to P. ultimum were planted.
Three replicates of 280 lines were tested for response to inoculation with F. graminearum in a seedling assay planted in growth media. Inoculated plants were assigned a root necrosis score, the ratio of uninoculated to inoculated root length and root dry weight are being determined, results are being calculated and data recorded for analysis.

Objective 2:
Data from phenotyping for resistance to 1) Fusarium graminearum, 2) P. ultimum, and 3) P. irregulare in the seedling assay is being prepared for association analysis.

Objective 3:
A. We have measured expressional changes of candidate genes for RpsUN1 and RpsUN2, respectively, at different time points (2, 4, 8, 12, 24 hours after inoculation) by quantitative real-time PCR, and cloned the candidate genes for RpsUN1 and RpsUN2 into a plasmid vector, and confirmed that the integrated gene sequences were correct by sequencing. We have been in the process of transferring the plasmids with integrated sequences of RpsUN1 and RpsUN2 candidate genes into Williams, a soybean variety that is susceptible to Phytophthora sojae for resistance evaluation suing whole plant as well as the hairy root systems.

B. We are using a special Phytophthora sojae strain with a molecular (Green Florescence Protein) tag to inoculate soybean hairy roots produced using Williams. This system will be used to evaluate phenotypic variation between transgenic and non-transgenic hairy roots

C. We are continuing the backcrossing process assisted with marker assisted selection towards the introgression of RpsUN1 and RpsUN2 into four Purdue elite cultivars. The backcrossing progeny seeds are being planted in the greenhouse and the progeny seeds from backcrosses in the greenhouse will be planted in the filed in early June.

Objective 4:
Progeny seed of crosses of RpsUN1 and RpsUN2 into prebreeding lines and elite breeding lines was increased in Chilean winter nursery.

Update:
Progress Report
Objective 1:
Seed, seedling, and root rot caused by F. graminearum and P. ultimum var.ultimum was evaluated in experiments to complete phenotyping of all 280 ancestral lines for resistance to and P. irregulare seedling rot (three experiments until the beginning of June. Further phenotyping effort in the greenhouse was postponed from June to late August because of excessively high and erratic temperatures that occured during the summer in the greenhouse. This trial, phenotyping for resistance to F. gramearum and P.ultimum, was resumed in September and has just been completed. Plants are now being evaluated and assigned a root necrosis scores, and root lengths, and root dry weights are being determined and data recorded. After completing this phenotyping step (F. graminearum and P. ultimum var.ultimum), three reps of the 280 ancestral lines will be planted and inoculated with P. irregulare to phenotype these lines for resistance to seed rot in vitro and seedling, and root rot in growth media. The third replicate of the F. graminearum phenotyping is also being planted upon completion breakdown and evaluation of the P. ultimum and F. graminearum experiments.
Objective 2:
(Minnesota) Data from phenotyping for resistance to 1) Fusarium graminearum, 2) P. ultimum, and 3) P. irregulare in the seedling assay is being prepared for association analysis to identify markers associated with QTLS for resistance to the three pathogens. (Purdue) We have developed a protocol to use soybean hairy root transformation system to validate RpsUN1 and RpsUN2 candidate genes that have been pinpointed by a combination of fine-mapping and candidate gene expression analysis. We are in the process of evaluating resistance of the candidate genes. Hope to get the results by the end of the year.

Objective 3: We have continued the backcrossing process assisted with marker assisted selection towards the introgression of RpsUN1 and RpsUN2 into four Purdue elite cultivars. The backcrosses have been done in the field and progeny seeds have just been harvested. These seeds are being planted in greenhouse for one generation of self-pollination. We have also analyzed the haplotypes of the genomic regions harboring the two candidate genes using the SNP genotyping data available for the entire USDA soybean germ plasm collection.

Objective 4: Progeny seed of crosses of RpsUN1 and RpsUN2 into elite breeding lines has been harvested from crosses in Chilean winter nursery during the winter of 2016 and 2017, was planted in local plots during the summer of 2017, and is now at the F4 plant stage.

Final Project Results

Update:
Objective 1:
Phenotyping of 280 ancestral lines for resistance to F. gramearum, P.ultimum, and P. irregulare seed and seedling rot was completed, root necrosis, root length, root dry weight measured and recorded for all lines for all pathogens.
Objective 2:
(Minnesota) Data recorded, evaluated for quality and control of variability (such as removal of outliers), and organized into datasets for statistical analysis and association mapping to identify markers for resistance. (Purdue) RpsUN1 and RpsUN2 candidate genes pinpointed by a combination of fine-mapping and candidate gene expression analysis.
Objective 3:
Progeny seed from introgression of RpsUN1 and RpsUN2 into four Purdue elite cultivars harvested. These seeds were planted in greenhouse for one generation of self-pollination. We completed analysis of the haplotypes of the two candidate genes using the SNP genotyping data available for the entire USDA soybean germ plasm collection.
Objective 4:
(Minnesota) Progeny seed of crosses of RpsUN1 and RpsUN2 with early maturity elite breeding lines planted for winter increase in Chilean nursery.

Benefit to Soybean Farmers

Performance Metrics

Project Years