Isolation and characterization of a gene expressed during early embryo sac development in apomictic guinea grass (Panicum maximum)

A novel classification method, based on ovary length, for sampling different developmental stages of embryo sac formation in obligate sexual and facultatively apomictic genotypes of guinea grass (Panicum maximum) has been employed to make a cDNA library and to isolate a stage-specific cDNA clone, probably representing a full-length gene. This A2-134 cDNA, designated ASG-1 (Apomixis Specific Gene), was found to be expressed in flower buds of the apomictic but not of the sexual accession. Furthermore, the gene was also not expressed in buds of apomicts until completion of megasporogenesis but could only be found during a phase characterized by the appearance of aposporous initial cells (AICs) of the embryo sac. The appearance of these cells is strictly limited to apomictic genotypes. Sequence analysis revealed that the A2-134 cDNA (1,177 bp) codes for a protein of 305 amino acids with a molecular mass of 34.2 kDa The amino acid sequence of A2-134 is related to RD22, a seed-specific and drought-induced gene of Arabidopsis thaliana, to USP, an unknown seed protein precursor of Vicia faba, to a polygalaturonase 1 beta chain precursor (Polyg1) of Lycopersicon esculentum and to ADR6, an auxin down-regulated gene of Glycine max. Southern blot analysis showed a different hybridization pattern at the genomic level when A2-134 cDNA was hybridized with total DNAs isolated from leaves of sexual as well as apomictic plants, indicating that the gene (or related family members) exists in both types. The cDNA library and the ASG-1 gene should be valuable tools toward understanding the molecular mechanism of apomixis, particularly apospory, and toward the transfer of apomixis to important crops. To our knowledge, A2-134 represents the first case of the isolation and characterization of a cDNA related to apomictic embryo sac development.