番茄mir172的表达模式分析与功能验证-expression pattern analysis and functional verification of tomato mir 172.docx

摘要在植物的生长发育过程中，miRNAs对器官的发育和分类起到了重要的作用。 它可以通过翻 译抑制或者 降解 mRNA来调控 靶向基因的表 达。在植物 众多的 miRNA家族中，miR172是其中非常保守的一类家族，该家族成员通过翻译抑制或 降解mRNA，或者二者兼而有之的方式来进行调节。之前有报道表明，miR172的 靶向基因是AP2类乙烯响应因子。本研究中，我们在Micro-Tom（Solanum lycopersicum L. cv.）野生型番茄里超 表达和抑制表达miR172家族的两个前体：pre-miR172b 和pre-miR172Hom，之后 通过定量PCR分析野生型和转基因番茄中的miRNAs表达模式，从而确定miR172 的功能。此外，我们还预测了8个miR172的AP2类靶向基因，同时分析了这8个基 因在野生型和转基因番茄中的表达水平。定量PCR结果显示，在野生型番茄的营养期内，miR172表达水平在茎中最 高；进入生殖期后，其表达量在破色期呈最低状态，之后逐渐恢复，红果期又达 到了最高水平。这也解释了番茄中超表达miR172b和miR172Hom后，植物会出现 了叶片和茎变宽增厚的原因。随着番茄在生殖期时的靶标基因SlAP2a, SlAP2h, SlAP2b 和SlAP2c 表达水平的提高，超表达miR172b和miR172Hom会使果实呈现出 膨大及方向型缺损的表型。此外，根据本研究发现，野生型番茄的果实从营养期 到成熟期的过渡阶段要早于超表达转基因番茄。表型分析及表达水平分析显示，植物中miR172对花的发育及果实的形成具有 瞬时调控的作用。有趣的是，在某一特定时期，miR172对于某些靶向基因并不总 是进行负向调节，二者的功能也有重叠的时候。尤其是在植物的破色期及红果 期，这一现象特别明显。这也有待于我们在今后的工作中继续深入研究。关键词：miR172，AP2 类基因，定量 PCR，相变，果实发育ABSTRACTmiRNAs play vital role in plant growth and physiology as well as organ development and identities through regulating the expression of their target genes either by translational repression or by mRNA cleavage. miR172 family is one of the conserved miRNA families across plant species, the members of this family can function by translational repression, mRNA cleavage, or both. miR172 has been reported to target AP2-like ethylene-responsive transcription factors.In this study, the function of miR172 was determined in Solanum lycopersicum L. cv. Micro-Tom (WT) by studying the effects of over-expressing and underexpressig two precursors of miR172 family i.e. pre-miR172b and pre-miR172Hom on the plant development, as well as by analyzing the expression patterns of both miRNAs in WT and the transgenic lines via qRT-PCR. Moreover, eight AP2-like genes were predicted to be targets of miR172 in tomato, therefore the expression pattern of each gene was also analyzed via qRT-PCR in both WT plants as well as transgenic plants.The expression pattern analyses of miR172 in WT revealed that miR172 is highly expressed in the stems during the vegetative phase, while