Deric Schmierer1, Arnis Druka1, David Kudrna1, and Andris Kleinhofs1,2
1Dept. of Crop and Soil Sciences;2School of Molecular Biosciences,
Washington State University, Pullman, WA 99164
Several chlorina seedling mutants have been identified in barley (fch1-fch15) that are dispersed throughout the 7 chromosomes (Franckowiak, 1997). fch12 is a single recessive trait characterized by yellow leaves with green tips at the seedling stage. As the plant develops, the leaf color changes to pale green (Robertson and Deming, 1930). Based on recent efforts to map morphological traits, fch12 was placed on the short arm of chromosome 7H flanked by the mutant genes brh1 and wax (Franckowiak, 1997).
The genetic stock fch12 mutant in a Bowman background was crossed to cv. Steptoe and cv. Morex to create a mapping population. Initially, a very small population of 25 individuals was used. A larger population of 85 individuals was then created for higher resolution. In both cases, plants that expressed the fch12 phenotype were selected in the F2 generation from the Steptoe cross. DNA was extracted and subjected to RFLP analysis via Southern blot hybridization by standard protocols used in our lab.
In the initial fch12 mapping population of 25 individuals, both Wax (KFP027) and BCD130 co-segregated with the mutant locus. When KFP027 was hybridized to the larger population, it mapped 2.3cM proximal to fch12. BCD130 still co-segregated with the mutant locus. The rice cDNA probe S10068 gave a poor hybridization signal on barley genomic DNA. For this reason, the probe was hybridized to several Horduem vulgare cDNA libraries. Clones that gave a strong signal were isolated from the cv. Morex seedling root library. These clones were sequenced and compared back to the original probe by both sequence analysis and confirmation hybridization. One EST, matching the S10068 sequence with near identical homology, was chosen for further work. This new probe, ABC327, co-segregated with fch12. ABG320 and Prx1A were used as distal and proximal flanking probes, respectively (Fig. 1).
References
Franckowiak, J.D. 1997. Revised linkage maps for morphological markers in barley, Hordeum vulgare. BGN 26:9-22.
Robertson, D.W., and G.W. Deming. 1930. Genetic studies in barley. J. Herd. 21:283-288.