1: Improve environmental stress and disease resistance in tree fruit crops. 1.A. Identify and characterize sources of fire blight resistance for use in apple scion breeding programs. 1.B. Characterize expression patterns and sequence differences of select apple drought-responsive genes in Malus sieversii lines exhibiting high and low water use efficiency. 1.C. Utilize transcriptomic and high-throughput genetic screening approaches to identify CBF-regulated and other stress-regulated genes, and characterize their functional role in stress tolerance and dormancy using transgenics and field evaluation. 2: Develop an accelerated breeding system for new tree fruit crops utilizing transgenic early-flowering lines.
Abiotic and biotic stresses play a major role in determining the economic viability of fruit crop production and postharvest quality. A single fire blight epidemic can destroy an entire young orchard and unfavorable environmental conditions, such as freezing temperatures, as well as heat and drought stress can result in significant reductions in yield, quality, and tree longevity. The overall objective of this project is to utilize genomic and molecular approaches to identify genes that convey resistance to abiotic and biotic stress in fruit crops, identify genetic markers for disease and stress resistance that can be utilized by apple breeders in marker-assisted-breeding programs, and to develop a breeding system that will facilitate the incorporation of specific traits, especially from novel genetic resources, such as Malus sieversii, into advanced selections of breeding lines. Quantitative trail loci (QTLs) and molecular markers for fire blight resistance will be developed for resistance derived from Malus sieversii and ‘Splendour’ apple. Targeted genome sequencing of the promoters of select dehydration and water use efficient responsive genes will be applied to lines of xeric-adapted Malus sieversii. Promoter analysis will identify cis-elements known to affect gene expression. Methylation differences between lines during simulated drought will be evaluated to reveal potential targets for gene regulation. The contribution of the CBF (C-repeat binding factor) family members to cold hardiness, dormancy, and growth will be evaluated. An accelerated breeding system for apple will be developed utilizing transgenic early-flowering lines to facilitate rapid integration of important genetic traits from novel apple genotypes into advanced breeding lines. The proposed research will result in the identification of genes, molecular markers, and a breeding system that can be used to efficiently develop apple germplasm with increased resistance to biotic and abiotic stress.
Host plant resistance is one of the most effective and sustainable options for managing fire blight, a devastating disease of apple and pear. Since genetic diversity is often lost during crop domestication, accessions of Malus sieversii, the wild progenitor of the domesticated apple, represent a valuable resource for disease resistance. Nearly 200 accessions of Malus sieversii from a USDA-ARS collection were selected as potential sources of disease resistance for apple scion breeding. Based upon controlled challenges with the fire blight pathogen, 12 wild Malus sieversii accessions were identified as resistant to fire blight with resistance comparable to highly resistant Malus × robusta 5 and resistant 'Delicious'. Three of these M. sieversii accessions PI657116, GMAL3688.c and GMAL4002.k were crossed in FY2017 with ‘Pinova’ (PinataTM) to begin incorporating this resistance into apple breeding programs. Developing frost protection methods for flowering fruit trees during spring frost events is critical. Transgenic MDCBFx plants with appropriate controls are near-ready for planting as indicated in the milestone goals for FY2018. An inter-graft experiment with CBF-overexpressing scions is underway to examine whether light perception is important for graft-transmission of altered dormancy. Constructs of several MDCBFx genes and their promoters have been created and transformation efforts begun in the model plant system Arabidopsis in order to better understand how these genes and their promoters operate. Breeding new apple varieties takes 25-30 years due to its long juvenile phase. Rapid cycling breeding using transgenic apple with reduced juvenility has the potential to greatly accelerate apple breeding. Crosses of reduced juvenility 'Pinova' T1190 with fire blight and apple scab resistant 'Enterprise' and blue mold resistant Malus sieversii resulted in the production of 1,100 reduced juvenility seedlings that are being selected for disease resistance. Validation of blue mold resistance markers in apple utilizing rapid cycling breeding technology. There is a need to incorporate disease resistance from wild apple genotypes into higher-quality apple genotypes and validate the genetic markers identified for the resistance trait. Crosses between reduced juvenility apple lines and blue mold resistant genotypes were made and seed was collected, stratified, and planted. Plants carrying the reduced juvenility gene were planted and pollinated with pollen from an 'Enterprise' parent. The resulting fruit is being evaluated for blue mold resistance and the presence of blue mold resistance markers. This will validate the blue mold resistance markers and begin the process of incorporating the resistance into a higher quality genetic background. Validation of the blue mold resistance markers and the incorporation of blue mold resistance into a better genetic background will provide a novel resource to apple breeders and prevent the need to treat apple fruit with fungicides to control the disease.
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