CBF Gene Copy Number Variation in Wheat and Barley and its Association with Freezing Tolerance and Winter Hardiness

Freezing tolerance and winter hardiness are complex traits. Over the course of the last 100 years different genetic tools have been employed with cereals of the Triticeae to gain insight into the complex nature of these traits. Emerging from many independent studies are findings that large affects are contributed by two regions on group five chromosome homoeologs, now identified as Frost Resistance-1 (FR-1) and FR-2. The FR-2 locus encompasses a cluster of genes encoding at least 11 different C-Repeat Binding Factor (CBF) proteins while FR-1 exerts negative control over expression of the CBF genes at FR-2. Recently we determined the physical structure of colinear segments of the FR-H2 region from four barley cultivars. Two of these were from freezing-tolerant winter-hardy genotypes and two of these were from less freezing-tolerant nonwinter-hardy genotypes. A key finding was that the genomic region encompassing subsets of the genes at FR-H2 is amplified to multiple copies occurring in tandem in the freezing-tolerant winter-hardy genotypes whereas the colinear genomic regions in spring genotypes occur in single copy. Other data suggest increased copy numbers are associated with increased transcript levels of the other CBF genes at FR-H2. Taken together these data suggest a complex relationship between gene dosage and the effects of the locus as a whole. We are currently utilizing different genetic tools including unstructured barley populations, wheat chromosome substitution lines, Agrobacterium-mediated transformation, and chromatin immunoprecipitation to further our mechanistic insight into FR-2 and its relationship with FR-1.