We have developed an in vitro system whereby we can reproduce the self-incompatibility (SI) reactions ofP. rhoeas in pollen grown in vitro, using stigmatic extracts. This has enabled us to investigate a number of aspects of SI, which would otherwise be difficult. On the stigma side of the reaction, the in vitro system has enabled us to characterize and partially purify the stigmatic S-component, following S-specific activity. It has also enabled us to establish that, in contrast to the S-linked glycoprotein ofNicotiana alata, no detectable ribonuclease activity correlates with the presence of the functional stigmatic S-gene product in this species. Turning to look at the pollen side, we have used the in vitro system to study the metabolic events occurring in the pollen ofP. rhoeas as a consequence of the SI reaction. We have determined that it requires both de novo glyco-sylation and RNA transcription for full inhibition of pollen-tube growth during the SI reaction. Transcription products of pollen SI response genes, which are produced specificially in an incompatible reaction, have been identified. These pollen response genes have been cloned and are currently being characterized. Since the extracellular pollen-stigma interaction results directly in gene transcription in the pollen, it seems likely that a signal transduction mechanism may be operating in the SI response. The in vitro system has allowed us to begin to investigate this possibility. We have detected rapid and transient phosphorylation of certain pollen proteins, together with changes in phosphatase activity during the SI reaction. These studies provide evidence for a role for signal transduction in the SI reaction. Thus, our in vitro system has enabled us to begin to examine, not only stigma and pollen components, but also the interaction between them in the SI reaction.