Developing rice endosperm cells display two distinct rough endoplasmic reticula (ER), cisternal ER (C-ER) and protein body ER (PB-ER), the latter delimiting the prolamine protein bodies. These ER membranes are utilized for the simultaneous synthesis of glutelin and prolamine storage proteins, which are subsequently routed into separate protein bodies. We demonstrate by blot hybridization, and by visualization of the spatial distributions and densities of these transcripts in endosperm cells via high resolution in situ hybridization analysis, that prolamine transcripts are associated primarily with the PB-ER, while glutelin mRNAs are enriched on the C-ER. The results suggest that the initial targeting process of these storage proteins into distinct protein bodies is the segregation of their transcripts on the ER membranes.