Sequencing of the Triticum monococcum hardness locus reveals good microcolinearity with rice

The Hardness ( Ha) locus on chromosome 5D is the main determinant of grain texture in hexaploid wheat. The related genes Puroindoline-a and -b ( Pina-D1, Pinb-D1) and Grain Softness Protein ( Gsp-D1) are tightly linked at this locus. Mutations in the Pina-D1 and Pinb-D1 genes are associated with increased grain hardness. We report here the complete sequence of a 101-kb BAC clone from Triticum monococcum (A(m ) genome) which includes these three genes, and its comparison with the orthologous region in rice. The genes Gsp-A(m) 1, Pina-A(m) 1 and Pinb-A(m) 1 are separated by 37 kb and 32 kb, respectively, and are organized in the same transcriptional orientation. Four additional genes, including a pair of duplicated genes, were identified upstream of Gsp-A(m) 1 within a high-density gene island. These additional genes were found in the same order and orientation, and the same relative distances apart as similar genes previously annotated on rice chromosome 12. An interesting discovery was a small unannotated putative rice gene that was similar to the Gsp-A(m) 1 gene of T. monococcum (65% similarity at the protein level), and that was disposed in the same orientation, and located in the same position relative to the other orthologous genes. The high gene density observed in this BAC (1 gene per 14 kb) was expected for a distal chromosome region, but the level of microcolinearity with rice was higher than that reported in similar distal regions of other wheat chromosomes. Most of the BAC sequence (40%) was represented by repetitive elements, mainly concentrated in regions adjacent to the genes Pina-A(m) 1 and Pinb-A(m) 1. Rearrangements among these repetitive elements might provide an explanation for the frequent deletions observed at this locus in the genomes of the polyploid wheat species.