Source: UNIVERSITY OF NEBRASKA submitted to
GENOME MAPPING OF BUFFALOGRASS [BUCHLOEDACTYLOIDES (NUTT.) ENGLEM]
Sponsoring Institution
National Institute of Food and Agriculture
Project Status
TERMINATED
Funding Source
HATCH
Reporting Frequency
Annual
Accession No.
0219181
Grant No.
(N/A)
Project No.
NEB-22-339
Proposal No.
(N/A)
Multistate No.
(N/A)
Program Code
(N/A)
Project Start Date
Jul 1, 2009
Project End Date
Jun 30, 2014
Grant Year
(N/A)
Project Director
Lee, D.
Recipient Organization
UNIVERSITY OF NEBRASKA
(N/A)
LINCOLN,NE 68583
Performing Department
Agronomy & Horticulture
Non Technical Summary
Buffalograss is proving to be a valuable choice for homeowners and turf managers who prioritized environmental factors in the landscape. New cultivars that provide better choices for consumers result from the expensive and time consuming process of plant breeding. A major expense in the plant breeding process comes with the repeated observations needed to make selections for successful growth when environment varies. One major environmental variable for buffalograss growth is chinch bug. If breeders could identify genes or chromosome regions that contain the genes that help cultivars in their growth reaction to chinch bugs, they would be able to select plants and eliminate the expensive, repeated observations that are now needed in the field and greenhouse. Selecting plants based on the DNA that marks specific genes is called marker assisted plant breeding. Marker assisted plant breeding can only happen when the following discoveries are made: 1) What chromosome regions have DNA sequence variation 2) What is the most reliable and cost effect way to detect these variable regions 3) Where are these variable regions located on the plant's chromosomes 4) Which of these chromosome regions or genes are associated with variation in important traits Our buffalograss genome mapping project will follow these four steps. Here are the key strategies we will use: 1) We have a funded project that will generate the DNA sequence of tens of thousands of chromosome regions in buffalograss. 2) We will use computer analysis of this massive DNA sequence data set to identify one type of DNA marker called the simple sequence repeat or SSR. 3) We are working with a family of buffalograss plants produced from crossing two diploid parents. This family represents the simplest gene inheritance situation in buffalograss and will be our first gene mapping family. 4) We have evidence from small scale chinch bug reaction studies with this family and that genes which impact the buffalograss chinch bug interaction vary among family members so mapping genes with chinch bug reaction is possible.
Animal Health Component
(N/A)
Research Effort Categories
Basic
(N/A)
Applied
(N/A)
Developmental
(N/A)
Classification

Knowledge Area (KA)Subject of Investigation (SOI)Field of Science (FOS)Percent
2011620108050%
2111620108125%
2111620108025%
Knowledge Area
211 - Insects, Mites, and Other Arthropods Affecting Plants; 201 - Plant Genome, Genetics, and Genetic Mechanisms;

Subject Of Investigation
1620 - Warm season perennial grasses;

Field Of Science
1080 - Genetics; 1081 - Breeding;
Goals / Objectives
Our goal is to map geneome regions in buffalograss and to identify associations between genome regions and chinch bug-tolerance. We are initiating these studies in a diploid population of buffalograss established by the UNL buffalograss breeding team. Diploid buffalograss cultivars are not a breeding goal. Rather, this population will serve as a model for testing hypotheses in genome organization and gene expression. We have five major objectives: Objective 1: Discover simple sequence repeat (SSR) genetic markers in buffalograss using 300+ megabases of 454 generated sequence from buffalograss . Objective 2: Discover SSRs from repeat enriched genomic sequences. Objective 3: Discover exon priming/ intron containing (EPIC) genetic markers that target chinch bug responsive genes in buffalograss. Objective 4: Map markers into the ten linkage groups expected in this species. Objective 5: Characterize chinch bug reaction inheritance in this diploid buffalograss population and find associations between markers and loci controlling these chinch bug reaction loci.
Project Methods
1. Markers discovered by 454 sequencing: We will discover new SSR DNA markers for buffalograss based on sequence data that will soon be generated from a large, second generation sequencing project. Focused analysis of the 5-10% of longest sequence reads from a 454 sequencing project can reveal repeat sequences. Repeat discovery will be made via http://www.gramene.org/db/searches/ssrtool. Primers can be designed that target these repeats. Repeat containing sequences will be identified and primers designed. These will be tested with a core set of 6-10 diverse buffalograss genotypes, including the diploid parents of our mapping population. This will be completed by 2011. II. Aternative SSR Discovery: It is expected that SSR markers from this large sequence database will be a robust start. However, we have an alternative SSR discovery strategy. We will use a scaled version of the SSR discovery and assessment approach as outlined below. - Genomic DNA will be isolated and restricted with the restriction enzyme AluI. - Restricted DNA will be enriched for SSR repeats by hybridization to (CA)20 oligonucleotides bound to a selection fiber. - Enriched fragments are cloned into a sequencing vector and a random sample of clones from the library sequenced and assessed for repeat enrichment. Repeat discovery will be made via http://www.gramene.org/db/searches/ssrtool - If the library has the enrichment level of 80-90%, intensive sequencing of this library will be done. If enrichment is lower, a second or third library using different restriction enzymes will be assembled and sampled for enrichment. - The new sequence database of this specialty crop will be screened for clones that contain repeats and primers will be designed that target these repeats. - SSR primers will be tested for polymorphism detection among the diploid population parents and across buffalograss germplasm. - Our goal is to identify 200 SSR markers that can be used in mapping in our current diploid population. III. Markers targeting introns (EPIC) will also be explored if SSR discovery is not robust. IV Mapping: The diploid population will serve as the model population for linkage mapping. This population will initiate our linkage mapping work for both gene specific and SSR markers. Our goal will be to map at least 10 loci per linkage group to establish a framework map of both markers and genes. Markers will include buffalograss-specific SSRs discovered in objective 5 and non-species specific markers discovered from past and current work. Mapping will be done using MapMaker. (http://linkage.rockefeller.edu/soft/mapmaker/ ) V. Markers and Chinch bug reaction: Current work in assessment of the reaction of diploid progeny to chinch bug feeding indicates the nature of inheritance of this trait in our crop. Distribution of progeny into a highly resistant (HR) phenotype category is consistent with the hypothesis that the diploid parents were heterozygous for alleles that have an additive effect on chinch bug response. Expansion of this experiment to the full 200 member population is warranted.

Progress 07/01/09 to 06/30/14

Outputs
Target Audience: Nothing Reported Changes/Problems: Marker systems attempted that were based on non specific DNA sequences have been replaced with new marker system development based on sequence from this species. What opportunities for training and professional development has the project provided? The technical staff person is being trained in the development and application of markers on this species. How have the results been disseminated to communities of interest? Nothing Reported What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

Impacts
What was accomplished under these goals? The buffalograss genome map is still to be established. A new faculty member was hired to take over this project two years ago.

Publications


    Progress 10/01/12 to 09/30/13

    Outputs
    Target Audience: Nothing to report Changes/Problems: Nothing Reported What opportunities for training and professional development has the project provided? New marker system development for this species. How have the results been disseminated to communities of interest? Nothing Reported What do you plan to do during the next reporting period to accomplish the goals? Nothing Reported

    Impacts
    What was accomplished under these goals? AFLP markers used on the same parents and populations as those mapped with SSRs are being deployed. Results are pending.

    Publications

    • Type: Journal Articles Status: Accepted Year Published: 2013 Citation: Serba, D.D., Guelson, Abeyo, B.G., Amundsen, K, Lee, D. , Heng-Moss, T., Baenziger, P., Eskridge, K, and Shearman, R. (2013) Turfgrass performance of diploid buffalograss [Buchloe dactyloides (Nutt.) Engelm.] half-sib populations. 47(2) 185-188.


    Progress 10/01/11 to 09/30/12

    Outputs
    OUTPUTS: Manuscript "Turfgrass Performance of Diploid Buffalograss (Buchloe dactyloides (Nutt.) Engelm.) Half-Sib Populations" submitted to HortScience. This work represents that transitional research from this Hatch to a newly developed Hatch project led by Dr. Keenan Amundsen. PARTICIPANTS: This will be the final year for this Hatch project as a new research leader has developed a new Hatch project. TARGET AUDIENCES: Not relevant to this project. PROJECT MODIFICATIONS: This will be the final year for this Hatch project as a new research leader has developed a new Hatch project.

    Impacts
    The research paper submitted to Hortscience explored and established the utility of genome mapping in a model diploid population to provide a foundational map for the buffalograss genome.

    Publications

    • Serba, D.D, O. Gulsen, B.G. Abeyo, K.L. Amundsen, D.J. Lee, P.S. Baenziger, T.M. Heng-Moss, K.M. Eskridge and R.C. Shearman. 2012. Turfgrass performance of diploid buffalograss [Buchloe dactyloides (Nutt.) Engelm.] half-sib populations. HortScience 47(2):185.


    Progress 10/01/10 to 09/30/11

    Outputs
    OUTPUTS: Preliminary DNA analysis study on Phragmites was conducted. The goal was to determine if tissue samples you collected as part of your Phragmities field sampling work would provide suitable tissue for DNA analysis. She also was able to provide a first look at the use of standard PCR primers that target chloroplast DNA sequences. These primers have been demonstrated to target the PCR amplification of chloroplast DNA in regions that are between chloroplast genes. Between gene regions often have DNA sequence variation so her third goal was to determine if she could detect DNA sequence variation based on a quick assay for DNA cutting differences (restriction fragment length differences or RFLPs). PARTICIPANTS: Project transitioned to support the initial work on Phragmities genetic variation for Dr. Steve Young, new faculty member in Agronomy and Horticulture, UNL. TARGET AUDIENCES: Preliminary data sets the stage for grant proposals and for collaborations with colleagues who currently study Phragmities genetic variation. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

    Impacts
    Outcomes were as follows: 1) The samples your provided gave Carol good quality DNA for analysis. 2) Standard chloroplast DNA PCR worked with the 'universal' primers. 3) Some DNA sequence variation was detected.

    Publications

    • Serba, D., T. Heng-Moss, R. Shearman, B. Abeyo, S. Baenziger, and D. Lee. 2011. Evaluation of buffalograss genotypes and full-sibs for chinch bug resistance. Arthropod-Plant Interactions.


    Progress 10/01/09 to 09/30/10

    Outputs
    OUTPUTS: Presentation at Crop Science Society of America meetings. PARTICIPANTS: This project has been the Ph.D. training research thesis work for Desalgn Serba. TARGET AUDIENCES: Not relevant to this project. PROJECT MODIFICATIONS: Major change in project will involve leadership as the University of Nebraska-Lincoln fills a new faculty line in turf breeding. It is anticipated that this new faculty we either take over this hatch or the hatch will close as a new Hatch funded program led by this scientist is developed.

    Impacts
    Demonstrated the feasibility of mapping in a diploid population which should improve mapping success in the future.

    Publications

    • Gulsen, O., T. Eickhoff, T. Heng-Moss, R. Shearman, F. Baxendale, G. Sarath, and D. Lee. 2010. Characterization of Peroxidase Changes in Resistant and Susceptible Warm-Season Turfgrasses Challenged by Blissus occiduus Barber. Arthropod-Plant Interactions 4: 45-55.


    Progress 10/01/08 to 09/30/09

    Outputs
    OUTPUTS: A first draft of the buffalograss linkage map has been developed in a diploid population. PARTICIPANTS: Nothing significant to report during this reporting period. TARGET AUDIENCES: Nothing significant to report during this reporting period. PROJECT MODIFICATIONS: Nothing significant to report during this reporting period.

    Impacts
    This work demonstrates the application and limitations of current DNA marker systems, the utility of the diploid population to provide recombination-based mapping information and the need to develop markers based on buffalograss genome sequence.

    Publications

    • No publications reported this period