Genetically and Physically Anchored EST Resources for Barley Genomics

Close, T.J.¹, Wing R.², Kleinhofs, A.³, Wise, R.⁴

¹Department of Botany & Plant Sciences, University of California, Riverside, CA, 92521-0124, USA; timothy.close@ucr.edu

²Clemson University Genomics Institute, Clemson, SC, 29634, USA; rwing@mail.clemson.edu

³ Department of Crop and Soil Sciences, Washington State University, Pullman, WA, 99164-6420, USA; andyk@wsu.edu

⁴Corn Insects and Crop Genetics Research, USDA-ARS and Department of Plant Pathology, Iowa State University, Ames, IA, 50011-1020, USA; rpwise@iastate.edu

Complete sequencing of large genome plants such as barley (5,000 Mb) is not yet practical. An alternative and well established route to about 70% of the genes is through cDNA libraries and expressed sequence tag (EST) sequences. A 1999 USDA/NRI grant to Rod Wing (Clemson University) and co-investigators Timothy Close (University of California, Riverside), Andris Kleinhofs (Washington State University), and Roger Wise (USDA-ARS/Iowa State University) supports the production for the public of a considerable barley EST resource (http://www.genome.clemson.edu/projects/barley<). The specific objectives are to: 1) develop high quality cDNA libraries from important developmental stages and tissue types; 2) sequence the 5' end of 50,000 barley cDNAs and deposit these to the GenBank EST database (dbest), and 3) anchor prioritized EST's to barley BAC clones and the barley genetic linkage map. In addition, the investigators will make use of rice genome sequences to identify rice orthologs and derive additional synteny-based map information. The project builds principally on Morex barley, both by the production of new materials and through the use of the existing Morex 6.3 x BAC library (Yu et al., 2000) and doubled haploid (DH) mapping populations that include Morex as one parent. Other mapping populations that can be easily linked to a map produced from a Morex-derived population, by virtue of mapped simple sequence repeats (SSRs) or other shared markers, will also be included. The Oregon Wolfe Barley DH population, which carries a high level of genetic polymorphism and has been developed as a teaching tool (http://www.css.orst.edu/barley/wolfebar/wolfnew.htm), is an excellent example.

The cDNA libraries in this project include a range of vegetative and reproductive tissues, and include normal as well as several biotic and abiotic stress treatments. The list of libraries, summarizing results as of January 2001, is shown in Table I. As of this date ten new cDNA libraries had been processed, each yielding about 5,000 randomly chosen cDNAs for a total of approximately 50,000 high quality ESTs that were submitted to GenBank. Additional information about each library and additional lower quality sequences that were not submitted to GenBank are posted at http://www.genome.clemson.edu/~dorrie/barley.html. The next phase of the project involves the identification of BAC clones that match a prioritized list of 600 barley ESTs, and genetically mapping as many of these as possible using the Steptoe x Morex, Harrington x Morex, Dicktoo x Morex and Oregon Wolfe Barley Mapping populations.

The list of 600 ESTs chosen for this phase of the work will include cDNA-derived markers that have already been placed on barley genetic linkage maps, and a prioritized list of genes that are of high interest. A general call has been made for input to this prioritized list (http://www.css.orst.edu/barley/nabgmp/Barley600.htm). Efforts have begun to coordinate the BAC anchoring and genetic mapping with other barley genome projects in Germany, Finland, Australia, Japan, France, Canada and the UK. The many investigators in this worldwide effort share a common vision of establishing a comprehensive barley microarray for functional genomics. It is our hope and intention that the products of worldwide cooperation and coordination on these fundamental and shared objectives in barley genetics will be swiftly and fully revealed to the public. In so doing, all barley improvement programs that utilize genetics will in a small period of time become more able to accelerate their progress toward practical end-product objectives, while at the same time the advantages of barley as a model plant will become increasingly evident to a broad spectrum of researchers and those who support research.

Reference

Yu, Y., J.P. Tomkins, R. Waugh, D.A. Frisch, D. Kudrna, A. Kleinhofs, R.S. Brueggeman, G.J. Muehlbauer, R.P. Wise and  R.A. Wing. 2000. A bacterial artificial chromosome library for barley (Hordeum vulgare L.) and the identification of clones containing putative resistance genes. Theoretical and Applied Genetics 101: 1093-1099.