EST generation and analyses towards identifying female gametophyte-specific genes in Zea mays L

The embryo sac (female gametophyte) plays an important role in double fertilization. The female gametophyte is composed of four specific cell types: the synergids that attract pollen tubes, the egg cell and central cell which are fusion partners for the two sperm cells, and the antipodal cells whose function is unknown. As a resource for gene discovery and to help identify genes exhibiting cell-specific expression patterns, we constructed cDNA libraries from female gametophytes and from egg cells of maize and sequenced more than 8,500 ESTs. These libraries represent diverse transcripts, potentially corresponding to 3,850 genes (contigs and singletons) from the female gametophyte and 963 genes (contigs and singletons) from the egg cell. In each collection, 16% of the contigs/singletons have no matches in databases and 3-5% encode hypothetical proteins; novel hypothetical proteins (not found within the female gametophyte contigs) were identified among the egg cell contigs. We examined 65 contigs by RT-PCR and 19 genes that were potentially female gametophyte-specific were identified. We used in situ hybridization to determine expression specificity for seven genes: one transcript was expressed both in the egg cell and in the central cell, one was expressed in the egg cell and synergids, two were expressed in the central cell, two were expressed in the synergids, and one was expressed in the central cell and the synergids. Four of these encode small, potentially secreted peptides that are dissimilar except for a conserved triple cysteine motif near their C-terminus. These EST resources should prove useful for identifying female gametophyte or cell-specific genes.