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Plant Genetic Resource Management, Preservation, Characterization and Utilization

Objective

<p>Objective 3: Evaluate priority crop core subsets and other selected germplasm with morphological descriptors, and key agronomic or horticultural traits, such as general adaptation, phenology, and growth potential. Identify accessions with desirable economical traits for multiple location tests and potential release to broaden the genetic base of breeding gene pools. </p>
<p>Objective 4: Apply molecular marker techniques to assess diversity, detect duplicated accessions, identify taxa that were difficult to classify with morphological characteristics and associated DNA polymorphism with variations of important economical traits in selected crops.</p>

More information

<p>NON-TECHNICAL SUMMARY: <br/>With the advent of molecular linkage maps valuable crop traits can be positioned on chromosomes with DNA-based markers. Knowledge of map position and closely linked DNA-based markers (tagging) can facilitate tracking and combining traits from different sources. Molecular markers, which function independent of the environment, allow for rapid selection of lines carrying the desired marker(s). One area of application is marker-assisted selection (MAS). MAS allows for the stacking, or pyramiding, of multiple disease resistance genes. These attributes directly benefit plant breeders through indirect detection of valuable traits, rapid high-throughput screening, and the ability to track multiple loci at once, thereby shortening the time to incorporate a new trait(s). Further benefits are gained through the detection of specific alleles, leading to
highly specific, easy-to-apply assays with increased resolution. A better understanding of crop genetics at the molecular level is an important component for further progress in food safety, nutrition, quality and yield.
<p>APPROACH: <br/>Select individuals from accessions with desirable traits such as disease resistance and abiotic stress tolerances during evaluation. After confirming the expression of the potential useful traits, the selected accessions will be promoted to the public. The association of molecular markers with a specific trait, qualitative or quantitative, provides an indirect method by which to track the trait independent of the environment. The goal of this objective is to contribute to the identification and development of diagnostic markers in potatoes and wheat. For traits with existing diagnostic assays it may be necessary to conduct proof-of-concept research to verify marker-trait associations when dealing with novel germplasm. A critical requirement is the localization of a trait on a molecular linkage map, followed by fine mapping or identification of a
tightly linked molecular marker. Research is also proposed for the development of laboratory friendly assays with the necessary attributes for diagnostic evaluation. More specialized marker development includes the design of muli-allelic (greater than 2 alleles) assays for polyploidy genotyping.

<p>PROGRESS:2013/01 TO 2013/09
<p>Target Audience: <br/>The target audience for the PLRV resistance research reported is primarily potato breeders, however this work indirectly will benefit potato growers and the larger potato industry. Target audiences for potato marker dosage research are potato breeders and other potato researchers. Target audiences for development of a new IMI diagnostic assay are wheat breeders and the wheat industry. Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? Attend and participate in the American Society of Plant Biologists meeting in Providence Rhode Island, July 2013. At this conference a poster was presented and numerous seminars attended on the latest research in plant science. Graduate student Ian Fullmer attended and participated in the annual meeting of the Potato
Association of America in Quebec, Canada. How have the results been disseminated to communities of interest? A talk was presented at the Potato Association of America conference in Quebec, Canada titled Molecular Characterization of Vacuolar Acid Invertase in a Population of Potatoes which shows Phenotypic Segregation for Fry Color in July 2013. This talk reported on the application of a pyrosequencing assay to quantify different alleles of vacuolar acid invertase in potato. A poster was presented at the Project Director Meeting for Plant Biology in Washington, D.C. May 2013. This reported progress in developing markers to track resistance to potato leafroll virus in potato. What do you plan to do during the next reporting period to accomplish the goals? Once BAC sequence data is available, alignment analysis will help identify the fewest possible contigs for the sequenced BACs. This
will form a physical map on which the associated molecular markers can be placed and surrounding genes identified. Depending of the extent of sequence coverage, additional BACs may be identified for sequencing. Sequence from S. etuberosum will be evaluated for evolutionary change when compared to potato and tomato sequence from homologous regions on chromosome 4. Additional markers can be developed and evaluated for linking to Rlretb. Sequence data from linked CAPS markers will be used to develop assays for quantifying alleles dosage in potato. Specifically sequence data from markers linked to potato genes Rysto and Rx1/Gpa2. Following assays design reaction conditions will be optimized and the assay tested against segregating progeny in a number of different potato populations.

<p>PROGRESS: 2012/01/01 TO 2012/12/31
<p>OUTPUTS: <br/>Research related to PLRV resistance has focused on Rlretb, a resistance locus from Solanum etuberosum mapped to potato chromosome 4. Previously indentified molecular markers were applied to the 109 genotypes from two backcross 4 populations and scored. A linkage map was generated using Map Manager. A BAC library generated from 109 selfed S. etuberosum seedlings was screened with molecular markers tightly linked to Rlretb. This screen identified 33 BACs which were subsequently submitted for sequencing. All identified BACs were screened with available molecular markers, thereby generating a series of overlapping BACs based on shared molecular markers. Additional efforts are being made to develop and characterize new molecular markers closer to Rlretb for use in marker assisted breeding. Research was initiated to develop
and evaluate diagnostic assays, using PCR and pyrosequencing, to replace existing PCR protocols for four potato resistance genes including Ryadg, Rysto, Rx1 and Gpa2. The goal of this work is to develop assays that can quantify resistance allele copy number in tetraploid potato cultivars. Initial work included reproducing previous reports, focusing on CAPS markers closely linked to the resistance genes. PCR fragments from resistant and susceptible lines were cloned and sequenced to determine sequence variation at the CAPS loci. A consensus sequence was used to design PCR primers and pyrosequencing primers that could be used to determine resistance allele dosage. Currently the assay for Ryadg is being optimized prior to conducting pyrosequencing. Sequences are being analyzed to design assays for Rx1 and Gpa2 prior to assay development.
<p>PARTICIPANTS: The principal investigator for this
project was Joseph Kuhl. He directed this research providing necessary technical and scientific support. He organized the research, developed the procedures that would be applied and oversaw application and recovery of results. He analyzed and interpreted results from the project mentioned above. The principal support scientist was Margaret Dibble. She applied protocols and collected results. Data was prepared and presented to the principal investigator. She applied the necessary protocols for the molecular markers used in the PLRV resistance research. She also applied the PCR assay used in the cloning of the vacuolar acid invertase project and the preparation of DNA for sequencing. Additional work was provided by undergraduate and graduate students working on various components of the above research. TARGET AUDIENCES: The target audience for the PLRV resistance research reported above
is primarily potato breeders, however this work indirectly will benefit potato growers and the larger potato industry. Target audiences for development of a new IMI diagnostic assay are wheat breeders and the wheat industry. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

Investigators
Kuhl, Joseph C
Institution
University of Idaho
Start date
2012
End date
2014
Project number
IDA01473
Accession number
228667
Commodities