Self-incompatible Petunia inflata and a self-compatible cultivar of Petunia hybrida will be used as model systems in an ongoing study of the molecular, biochemical, and genetic bases of gametophytic self-incompatibility. The first objective is to provide direct evidence for the role of the S-gene (the gene encoding the pistil S-protein) in the recognition and rejection of self pollen. Two approaches will be taken: a) transgenic P. inflata plants obtained by introducing S1-gene into S3S3 genotype, and S3-gene into S1S1 genotype, will be tested to see whether the ability to reject pollen bearing S1-allele and S3-allele respectively has been conferred by the transgenes; b) antisense S1-gene will be introduced into P. inflata plants with S1S3 genotype to see whether the ability to reject S1-bearing pollen is annulled in the transgenic plants. The second objective is to determine the regulatory sequences for the pistil-specific expression of the S- gene and rnx2 gene (encoding RNase X2) by examining the expression of a series of promoter-GUS constructs via transient assays and in stable transformants. The third objective is to identify and characterize other components that participate in self- incompatibility interactions. PCR techniques will be used to investigate whether the S-gene is expressed in pollen during its growth in situ. Chromosome walking experiments will be used to identify sequences linked to the S-gene that might encode other components of the self-incompatibility system. An approach based on immunoprecipitation will be used to examine whether pollinated pistils contain any proteins that interact with S-proteins. These experiments may lead to identification of the long sought pollen component of the S-locus. The fourth and final objective is to determine the genetic basis for the breakdown of self- incompatibility in the self-compatible cultivar of P. hybrida. Genetic crosses have been designed to test a model implicating a modifier locus in the modulation of S-alleles activity in self- incompatibility interactions.
