Location: Innovative Fruit Production, Improvement, and Protection
2020 Annual Report
Objectives
Objective 1: Develop elite breeding parents for apple containing both disease resistance alleles to blue mold, apple scab, and fire blight, and desirable fruit quality alleles using rapid cycle breeding technology. [NP301, C1, PS1B]
Objective 2: Utilize Malus sieversii (an apple progenitor species) and a forward overexpressing system (FOX-hunting) to identify and functionally analyze apple genes conferring disease and environmental stress resistance. [NP301, C3, PS3A]
Objective 3: Utilize CRISPR technology to improve disease resistance or environmental stress tolerance in 'Royal Gala'. [NP301, C1, PS1B]
Approach
Environmental stress and outbreaks of disease can severely reduce apple fruit production. New cultivars with improved stress and disease resistance will be needed in order for the U.S. apple industry to remain competitive. Rapid cycle breeding will be used to combine multiple disease resistance alleles for fire blight, apple scab and blue mold resistance within genetic backgrounds that are suitable for use in apple breeding programs. DNA-informed breeding methods, rather than conventional phenotyping methods, will be used to predict individuals with multiple sources of disease resistance and fruit quality traits. The wild apple species, Malus sieversii, is considered to represent a reservoir of disease and stress resistance genes, as well as a source of novel, quality traits (flavour, texture, etc.). A Full-length cDNA Over-eXpressing (FOX) Gene Hunting system will be used to conduct high-throughput screening of genes in Malus sieversii – PI613981 (disease and drought tolerant elite genotype) that are associated with freezing, drought, and salt tolerance. Once vetted, the identified genes will be used to produce apple genotypes of ‘Royal Gala’ with improved stress tolerance. Lastly, several potential genes regulating time of bud break will be characterized in transgenic and non-transgenic apple lines to confirm their role in the regulation of bud break. Key regulatory motifs in the promoters of these genes will be modified using CRISPR/Cas9 technology to produce genotypes with delayed bud break in order to adapt apple genotypes to erratic spring weather patterns.
Progress Report
This is a final report for project 8080-21000-026-00D due to a redirection to project 8080-21000-029-00D.
The rapid cycle breeding system for apple that utilizes transgenic, early-flowering trees has been implemented and used to generate genotypes with pyramided resistance (two unique and independent sources of fire blight resistance) to fire blight and blue mold resistance. Further crosses have been made to dilute out the unwanted Malus sieversii (wild apple) genetic background and select for good fruit quality traits, while maintaining fire blight and blue mold resistance traits. All screening and selection is being based on the use of molecular markers. The project was awarded an ARS Innovation Fund Award to develop advanced apple breeding lines for distribution to apple breeders.
Malus sieversii, the progenitor of the modern apple, is considered a highly valuable resource for biotic and abiotic stress resistance genes. A highly-valued M. sieversii selection, PI613981, has been used to create over 12,000 lines of Arabidopsis, each carrying a unique gene from M. sieversii. The lines were sequenced to identify each inserted gene. Phenotyping assays were developed to screen the lines for cold, salt, and drought tolerance, as well as developmental regulation (architecture and general morphology). Thirteen hundred of the apple FOX lines were screened for cold tolerance. All the lines have been cataloged and preserved for future distribution to interested researchers.
Developing frost protection methods for flowering fruit trees during spring frost events is critical. Poplar lines overexpressing the native poplar EBB1 (Early BudBreak1) gene were shown to break dormancy early. Transgenic MdEBB (apple EBB) trees have been created along with trees overexpressing the poplar EBB1 gene. Trees are growing in the greenhouse and are being evaluated as to changes in dormancy and growth habit. A better understanding of dormancy and delayed budbreak via directed gene editing in these trees is anticipated.
Accomplishments
Review Publications
Luo, F., Van De Weg, E., Vanderzande, S., Norelli, J.L., Flachowsky, H., Hanke, V., Peace, C. 2019. Elucidating the genetic background of the early-flowering transgenic genetic stock T1190 with a high-density SNP array. Molecular Breeding. https://doi.org/10.1007/s11032-019-0929-z.
Luo, F., Norelli, J.L., Howard, N., Wisniewski, M.E., Flachowsky, H., Viola-Hanke, M., Peace, C. 2020. Introgressing blue mold resistance into elite apple germplasm by rapid cycle breeding and foreground and background DNA-informed selection. Tree Genetics and Genomes. https://doi.org/10.1007/s11295-020-1419-5.
