Cloning of genes preferentially expressed in early stage of parthenocarpic and pollinated fruit and stabilization of fruit set

本项目旨在明确孤雌结实和坐果的优先相关基因，以稳定其稳定性。荧光显微镜观察到，茄子花授粉后花粉管伸长，在授粉后1.5 ~ 2天内，花粉管尖端到达子房胚珠。因此，在开花期阉割后设置授粉、不授粉和不授粉+生长素处理3个实验条件。每个样本在生长2或3天后提取rna。对于授粉后2 d的表达，通过授粉果与未授粉果之间的相减，制备了相减cDNA文库。我们随机挑选了384个cDNA克隆，并将它们定位在膜上进行大阵列分析。选择了32个在授粉果实中表达高于未授粉果实的候选cDNA克隆作为探针进行northern blot分析。结果表明，授粉果实的mRNA杂交信号强于未授粉果实的mRNA杂交信号。与P450、Unknown蛋白和Hypothetical蛋白相似的3个基因的mrka在花后2 ~ 4天在授粉和生长素处理的果实中特异性表达，然后迅速减少。另一方面，在授粉后3天的表达中，有10个cDNA克隆优先表达，其中包含组蛋白、花心蛋白、多酚氧化酶、细胞壁结构蛋白、细胞分裂相关蛋白、分子伴侣子等。然而，几乎所有mrna在3天后仍能清晰表达。因此，这3个基因在2天内的表达与坐果形成有关。在甜椒果实中，也用同样的方法分离到了7个EST基因，这些基因在授粉后2天优先在茄子果实中表达。胚体花2主要通过传粉方式表达，具有调控开花的功能。少

The purpose of this project is to make clear the genes preferentially relating to parthenocarpy and fruit setting for its stabilization.It was obserbed using fluorescence microscopy that pollen tube elongated after pollination of eggplant flower and the tip reached the ovules in ovary within 1.5 to 2 days after pollination. Therefore, three experimental conditions of pollination, unpollination, and unpollination + auxin treatments were set after emasculation on the flower opening. And RNAs from each sample were extracted after 2 or 3 days growth. Regarding the expression on 2 days after pollination, subtracted cDNA library was prepared by the subtraction between pollinated and unpollinated fruit. We picked up randumly 384 cDNA clones and spotted them on the membrane for macroarray assay. Thirty two candidates of cDNA clones for genes expressed higher in pollinated fruit than unpollinated one were selected and used as probes for Nothern blot analysis. Eleven of these 32 cDNA clones show … More ed that hybridization signals with mRNA from pollinated fruit were stronger ttian that from unpollinated fruit. mRKAs of three genes having similarity to P450, Unknown protein, and Hypothetical protein were specifically expressed in both pollinated and auxin treated fruit from 2 to 4 days after anthesis and then reduced rapidly. On the other hand, regarding the expression on 3 days after pollination, there were 10 cDNA clones expressed preferentially, in which histone, carleticulin, polyphenol oxidase, cell wall structural protein, cell division relating protein, molecular chaperon were contained. However, almost all mRNAs continued to be expressed clearly ever after 3 days. Thus, it was suggested that the 3 genes expressed on 2 days related to the initiation of fruit set.In sweet pepper fruit, also, 7 EST genes preferentially expressed in pollinated fruit on 2 days after pollination were isolated using the same way to eggplant fruit. Embryomic flower 2, which functions the regulation of flowering, was prominently expressed by pollination. Less