ISOLATION OF MOLECULAR MARKERS FROM THE BARLEY ENDOSPERM COENOCYTE AND THE SURROUNDING NUCELLUS CELL-LAYERS

The cereal endosperm develops from a coenocyte to a cellular storage o rgan through formation of nucleo-cytoplasmic domains and cell wall dep osition in the interzones between these domains. During its early stag es, the endosperm develops in close contact with nucellus, the sporoph ytic tissue which gives rise to the megagametophyte. Owing to the posi tioning of the two tissues deeply within the ovary, neither cell types have been easily accessible for molecular studies. In this paper we r eport for the first time the cloning of molecular markers for the barl ey endosperm coenocyte and the nucellus. The novel END1 and NUC1 cDNAs were isolated by differential screening of a cDNA library from 5 DAP (days after pollination) ovaries using a positive probe from hand-diss ected embryo sacs with adhering nucellus and testa cell layers, and a negative probe from pericarp. In situ and northern blot hybridization data show that END1 transcripts are asymmetrically distributed in the endosperm coenocyte limited to an area over the nucellar projection. I n the cellular endosperm, END1 transcripts are present in modified ale urone cells and a few layers of ventral starchy endosperm cells. The s econd clone, NUC1, hybridizes to transcripts in the nucellus before fe rtilization and in autolyzing nucellus cells after fertilization. At l ater stages, after the disappearance of nucellus, NUC1 transcripts are present in the nucellar epidermis and in the lateral cells of the nuc ellar projection. This work provide tools for future elucidation of th e genes specifying endosperm histogenesis.