Analyses of molecular genetic mechanism involved in the sporophytic self-incompatibility of Ipomoea.

自交不亲和（SI）是一种阻止自交受精从而促进异交以维持植物群体遗传多样性的遗传系统。SI系统通过雌蕊识别自性花粉和非自性花粉而排斥自性花粉来工作。在旋花科植物中，几个品种的易普莫亚（Ipomoea）具有孢子体型的SI，由一个单一的多等位基因s位点调控。本研究旨在鉴定与Ipomoea自花花粉识别相关的s位点基因，并揭示自花花粉识别系统的分子机制。研究结果总结如下。(1)构建了s位点附近DNA标记的精细遗传图谱。(2)采用基于图谱的定位克隆策略，从细菌人工染色体（BAC）和3种S单倍型的cosmid文库中筛选出覆盖S位点的基因组克隆。(3)对分离克隆的序列分析表明，S-座被划入S单倍型特异性发散区（SDRs），大小在30 ~ 95 kb之间。(4)通过对mrna的Northern blot和RT-PCR分析，确定了3个柱头特异性基因（SE1、SE2、SEA）和1个花药特异性基因（AB2）分别编码雄蕊和花粉决定因子。(5)用RNAi构建的柱头特异性基因引入的转化子在SI表型上没有变化，表明柱头特异性基因可能具有互补作用。(6)本研究表明，这些柱头特异性候选基因与迄今为止报道的其他植物的s位点基因没有同源性，表明Ipomoea植物的SI系统具有独特的分子机制。

Self-incompatibility (SI) is a genetic system that prevents self-fertilization and thereby promotes out-crossing to maintain genetic diversity in plant populations. The SI system works through the rejection of self-pollen following the recognition of self and non-self pollen by the pistil. In the plant family, Convolvulaceae, several species of Ipomoea possess a sporophytic type of SI that is regulated by a single multiallelic S-locus. The present study was performed to identify the S-locus genes involved in the self-pollen recognition and to reveal molecular mechanism of the SI system in Ipomoea. Research results obtained are summarized as follows. (1) Fine genetic map of DNA markers was constructed in the vicinity of the S-locus. (2) Using a map-based positional cloning strategy, several genomic clones covering the S-locus were screened from bacterial artificial chromosome (BAC) and cosmid libraries of three S haplotypes. (3) Sequence analysis of the isolated clones showed that the S-loci were delimited to the S haplotype-specific divergent regions (SDRs) spanning 30 to 95 kb in size. (4) From gene expression analyses by Northern blot and RT-PCR of mRNAs, three stigma-specific genes (SE1, SE2, SEA) and an anther-specific gene (AB2) in the SDRs were identified as candidates encoding pistil and pollen determinants of the SI, respectively. (5) Transformants introduced with RNAi constructs of the stigma-specific genes showed no changes in the SI phenotypes, indicating that the stigma-specific genes might be functioned as complementary. (6) The present study revealed that these stigma specific candidates showed no homologies with the S-locus genes reported to date from other plants, and suggests that a unique molecular mechanism is involved in the SI system of Ipomoea plants.