黄瓜microRNA171的克隆与功能分析

Cloning and Functional Analysis of microRNA171 in Cucumber

为了解microRNA171(miR171)对黄瓜的调控作用,从华北型黄瓜‘北京截头’中克隆了Csa-miR171家族的4个miRNA前体序列Csa-miR171a、Csa-miR171b、Csa-miR171c、Csa-miR171d,对前体序列进行比对与进化分析,预测成熟序列的靶基因并进一步开展了拟南芥遗传转化功能研究。结果表明,4个前体共剪切出两种成熟序列Csa-miR171a和Csa-miR171b,系统进化显示,Csa-miR171a、Csa-miR171b、Csa-miR171c、Csa-miR171d分别与甜瓜Cme-miR171e、Cme-miR171c、Cme-miR171a、Cme-miR171b同源性最近,与拟南芥、水稻、番茄等较远;Csa-miR171a、Csa-miR171b成熟序列主要靶向GRAS转录因子蛋白家族基因,拟南芥中过表达Csa-miR171a前体改变了叶片形态、颜色、抽薹时间、根、果实、花器官形态等多个表型性状。

To investigate the function of microRNA171(miR171)in cucumber,four precursors Csa-miR171a,Csa-miR171b,Csa-miR171c,Csa-miR171d of Csa-miR171 family genes were cloned in the northern Chinese cucumber‘Beijingjietou’and followed by sequence alignment and phylogenetic analysis.Target genes of the mature sequence of Csa-miR171 were predicted and functional analysis was performed in Arabidopsis by genetic transformation.The results showed that the two mature sequences,Csa-miR171a and Csa-miR171b were cleaved from the four Csa-miR171 precursors.Phylogenetic analysis showed that Csa-miR171a,Csa-miR171 b,Csa-miR171c,Csa-miR171d were closer to Cme-miR171e,Cme-miR171c,Cme-miR171a,Cme-miR171b in melon than those in Arabidopsis,rice and tomato,etc.GRAS transcription factor family was predicted to be the main target gene of Csa-miR171.Overexpression of Csa-miR171a in Arabidopsis significantly affected the phenotypes including leaf morphology and leaf color,blooming time,root,fruit and floral morphology.