烟草(Nicotiana tabacum)生长素响应基因NtIAA27 的克隆及功能分析

为了研究内源生长素信号路径对烟草烟碱合成的调控,本研究根据烟草基因组数据和PCR扩增获得烟草生长素响应蛋白NtIAA27基因RNAi干扰片段180 bp,将该基因180 bp的片段克隆至RNAi载体pHellsgate12中,并通过叶盘法将该基因转入烟草K326中。经过实时定量PCR检测获得两株敲除效率较高的转基因植株,对这两株转基因株系及非转基因材料进行烟碱含量测定,结果表明与非转基因植株相比,RNAi转基因植株的烟碱含量升高,表明内源生长素信号路径可以负调控烟草烟碱含量,本研究将为培育不同烟碱含量的烟草品种提供有价值的信息。

Cloning and Functional Analysis of an Auxin Responsive Gene NtIAA27 from Tobacco

To investigate regulation of the endogenous auxin signaling pathway on the biosynthesis of tobacco nicotine,a 180 bp DNA fragment of auxin responsive protein NtIAA27 gene was obtained by PCR according to the tobacco genome databank in the present study.The fragment was cloned into pHellsgate12 to construct an RNAi expression vector and the NtIAA27-RNAi vector was transformed into tobacco K326 by the Agrobacterium-mediated leaf disk method.Two independent transgenic lines with high knockout efficiency were confirmed by real-time PCR.The assay of nicotine contents in non-transgenic plants and transgenic plants showed that the transgenic plants of NtIAA27-RNAi had higher nicotine content than the non-transgenic plants,which implied that the endogenous auxin signaling pathway could modulate the nicotine content.The present study provides helpful information for breeding tobaccos with various nicotines.