Molecular genetical analysis of the two locus self incompatibility in the grasses (Poaceae)

禾本科植物的自交不亲和性是由两个互不连锁的基因座S和Z控制的，本课题的目的是建立野生大麦球茎大麦自交不亲和性的遗传基础，并克隆S的自交不亲和性基因。基本的遗传分析包括植物品系杂交试验、SI基因类型的确定、大作图群体的建立、与SI基因座紧密连锁的DNA标记的鉴定等。针对S基因的克隆，采用基于AFLP的基因指纹图谱方法从花药和雌蕊中寻找S基因型特异的转录本。通过AMF分析，在雌蕊和花药转录本中都检测到了数十个S特有的片段。对克隆的片段进行限制性片段长度多态性分析，验证其与S基因座的连锁关系。到目前为止，已有几个克隆被确认为与S基因座的紧密连锁。利用与S连锁的无性系构建了S基因座周围的详细遗传图谱。S基因座与部分克隆间未见重组，表明这些克隆是S基因的候选基因。应对候选克隆的表达模式和序列多态性进行详细分析，以确定它们是否是S基因。还有必要建立球茎海棠的转化体系，以证明候选基因是S专一性的决定因素

The self-incompatibility (SI) in the grasses (Poaceae) is gametophytically controlled by two unlinked loci, Sand Z, The aim of this project is to establish the genetic basis of the SI in wild barley, Hordeum bulbosum (2x) and to clone the SI gene(s). The basic genetical analysis involves crossing experiments of plant lines, determination of SI genotypes, production of large mapping populations, identification of DNA markers closely linked to the SI loci, etc. Toward the,, cloning of the S gene, the AFLP-based mRNA fingerprinting (AMF) method was employed to search S genotype-specific transcripts from pistils and anthers at anthesis. By the AMF analysis, dozens of S specific fragments were detected from both the pistil and anther transcripts. The fragments were cloned and subjected to the RFLP analysis to examine linkage to the S locus. Several clones have so far been confirmed for the close linkage to the S locus. The S-linked clones were used to construct a detailed genetic map around the S locus. No recombination was found between the S locus and some of the clones, indicating that those clones are candidates of the S gene. The expression pattern and sequence polymorphism of the candidate clones should be analyzed in detail to examine whether they are the S gene or not. It is also necessary to develop the transformation system for H bulbosum to demonstrate that the candidate gene is the determinant of S specificity