2010 Annual Report
1a.Objectives (from AD-416)
Utilize functional genomic/proteomic approaches to identify genes or proteins in fruit crops that confer resistance or susceptibility to freezing or water stress and fire blight. Develop and evaluate new transgenic technologies needed to control gene flow and expression in transgenic apple trees. Evaluate genetically engineered rootstocks as (1) a means of enhancing resistance to diseases and (2) a means of indirectly influencing gene expression in the scion for the improvement of performance or quality.
1b.Approach (from AD-416)
Transcript profiling will be used to identify genes associated with environmental stress, fire blight resistance, and susceptibility, while proteomic approaches will be used to better understand the relationship between gene expression and cognate protein levels. RNAi-induced gene siliencing will be used to elucidate the role of specific candidate genes in resistance and adaptation. The ability to use genetically-engineered rootstocks of apple for scion trait modification will be explored by investigating graft-transmissible gene silencing. The use of floral-specific promoters to confine and regulate the expression of dehydrin genes (responsible for cold and dehydration tolerance) to flowers of fruit crops in order to improve frost tolerance will also be investigated.
This project will terminate at the end of September, 2010. A bridge project will be in effect beginning October 1, 2010 until 2013 to allow for the merger of NP302 with NP301 and synchronization of OSQR evaluation of new NP301 Projects. New milestones have been developed and will be reported on in FY 2011. The bridge project will represent the continuation of some of the research initiated in the terminating project and the logical definition of new goals. Substantial results were obtained over the five years of the project. New genomic tools were developed to address critical problems of biotic and abiotic stress resistance in apple, and transformation technologies were developed to study the function of candidate genes involved in stress resistance. In collaboration with the University of Illinois, a 40,000 apple unigene microarray was developed for apple with applicability to other Rosaceae crop plants (peach, pear, strawberry). Using multiple genomic approaches, candidate genes were identified for fire blight resistance, as well as cold and drought resistance. A genetic marker for fire blight resistance was developed in collaboration with scientists from the New Zealand Institute for Plant and Food Research. Genes associated with drought response have been identified using subtractive/suppressive hybridization technology and are being further evaluated for their contribution to drought resistance. Complete families of dehydrin genes have been identified in both peach and apple, and their induction by low temperature and drought has been characterized. CBF transcription factors have been identified and characterized in both apple and peach, and overexpression and silencing studies have demonstrated their role in cold and drought resistance. The apple microarray has been used to identify fire blight inducible genes and genes regulated by the CBF transcription factor. One CBF gene has been demonstrated to regulate induction of dormancy in apple in response to short days. The application of genomic approaches has clearly been demonstrated to be of great value to identify candidate genes for biotic and abiotic stress resistance, as well as specific markers for fire blight resistance. Near the end of the project, an effort was initiated to develop a genetic map for Malus sieversii, a wild species of apple considered to be a progenitor of the domesticated apple and an excellent resource for new, economically important traits. Additionally, efforts have been initiated to develop transgenic early flowering lines of apple to use as a system to rapidly incorporate important traits from novel apple germplasm, such as Malus sieversii, into advanced breeding material. The mapping work with Malus sieversii and the development of early flowering lines of apple for efficient breeding are efforts that will continue in the bridge project.
Transgenic Apple Trees with enhanced Tolerance to Climate Change. Apple trees are not responsive to short days but rather enter into dormancy after exposure to low temperatures. This can make them more prone to injury from the erratic weather patterns predicted by climate change models. Extended warm temperatures in the fall, winter, or spring followed by unseasonable low freezing temperatures can cause severe injury and result in millions of dollars of crop loss. Transgenic apple plants have been created by ARS scientists at Kearneysville, WV that overexpress an AP2-transcription factor. The transgenic trees respond to short days as the cue to enter dormancy rather than just low temperature. This may make them more adaptable to climate change and unpredictable temperature changes, thereby preventing severe injury and crop loss.
Molecular DNA Marker for Fire Blight Resistance developed. The U.S. tree fruit industry has identified the implementation of marker assisted breeding programs as a priority in order to accelerate and improve efforts to generate commercially-desirable scion and rootstock cultivars. Fire blight is one of the most feared diseases for apple and pear growers because infections of young trees grown on susceptible rootstocks often lead to tree death. A DNA marker for fire blight resistance was developed by ARS scientists at Kearneysville, WV in collaboration with scientists at the New Zealand Institute for Plant and Food Research. The marker is based upon a specific receptor like protein gene (allele) found in a source of fire blight resistance, Malus ‘Robusta 5’, that is widely used in apple breeding programs. This gene was previously associated with fire blight resistance and the marker was detected in fire blight resistant progeny of ‘Robusta 5’ but not in susceptible progeny or other susceptible cultivars. After validation, the marker will be made available to U.S. tree fruit breeding programs to facilitate the development of fire blight resistant apple and pear cultivars by marker assisted breeding.
Malnoy, M., Boresjza-Wysocka, E., Norelli, J.L., Flaishman, M., Gidoni, D., Aldwinckle, H.S. 2010. Genetic transformation of apple (Malus x domestica) without use of a selectable marker gene. Tree Genetics and Genomes. 6:423-433.
Karlson, D., Byard, S., Wisniewski, M.E., Li, J. 2010. Interspecific analysis of xylem freezing responses in Acer and Betula. HortScience. 45:165-168.
Macarisin, D., Wisniewski, M.E., Bassett, C.L., Thannhauser, T.W. 2009. Proteomic analysis of B-aminobutyric acid priming and aba-induction of drought resistance in crabapple (Malus pumila): effect on general metabolism, the phenylpropanoid pathway and cell wall enzymes. Plant Cell and Environment. 32: 1612-1631.