Heat shock-inducible clonal analysis reveals the stepwise establishment of cell fate in the rice stem

The stem, consisting of nodes and internodes, is the shoot axis, which supports aboveground organs and connects them to roots. In contrast to other organs, developmental processes of the stem remain elusive, especially those initiating nodes and internodes. By introducing an intron into the Cre recombinase gene, we established a heat shock-inducible clonal analysis system in a single binary vector and applied it to the stem in the flag leaf phytomer of rice (Oryza sativa). With detailed characterizations of stem structure and development, we show that cell fate acquisition for each domain of the stem occurs stepwise. Cell fate for a single phytomer was established in the shoot apical meristem (SAM) by one plastochron before leaf initiation. Cells destined for the foot (nonelongating domain at the stem base) also started emerging before leaf initiation. Cell fate acquisition for the node began just before leaf initiation at the flank of the SAM, separating cell lineages for leaves and stems. Subsequently, cell fates for the axillary bud were established in early leaf primordia. Finally, cells committed to the internode emerged from, at most, a few cell tiers of the 12- to 25-cell stage stem epidermis. Thus, internode cell fate is established last during stem development. This study provides the groundwork to unveil underlying molecular mechanisms in stem development and a valuable tool for clonal analysis, which can be applied to various species.