Molecular Genetic Basis of Self-Incompatibility in Petunia

自交不亲和的矮牵牛和自交亲和的 矮牵牛将被用作正在进行的研究中的模型系统 配子体的分子、生物化学和遗传基础 自交不亲和性第一个目标是提供直接 S基因（编码雌蕊的基因）作用的证据 S-蛋白）在识别和排斥自花花粉中的作用。两 a）获得转基因P. inflata植物， 将S1基因导入S3 S3基因型，将S3基因导入S1 S1基因型， 基因型，将被测试，看看是否有能力拒绝 分别带有S_1和S_3等位基因的花粉， B）反义S1-基因将被 引入具有S1 S3基因型的胀囊青霉植物中，以观察是否 拒绝S1花粉的能力在 转基因植株第二个目标是确定 雌蕊特异性表达S- 基因和rnx 2基因（编码RNase X2）的表达 一系列启动子-GUS构建体的瞬时表达， 稳定的转化体。第三个目标是确定和 表征参与自- 不相容的相互作用PCR技术将用于 研究S基因是否在花粉发育过程中表达， 生长在原地。染色体行走实验将用于 确定与S基因相连的序列，这些序列可能编码其他 自交不亲和系统的组成部分。基础的办法 免疫沉淀将被用来检查是否授粉 雌蕊含有任何与S蛋白相互作用的蛋白质。这些 实验可能会鉴定出长期寻找的花粉 S-轨迹的组成部分。第四个也是最后一个目标是 确定自我崩溃的遗传基础， 自交不亲和品种的不亲和性。 遗传杂交被设计用来测试一个模型， 修饰基因座在自我调节S等位基因活性中的作用 不相容的相互作用

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.