人参AGO1基因的克隆及序列分析

Cloning and sequence analysis of AGO1 gene in Panax ginseng

目的:从人参叶片中克隆Argonaute 1(PgAGO1)的全长基因,并利用生物信息学方法进行分析。方法:根据人参EST序列设计特异性引物,采用RACE方法克隆PgAGO1的cDNA全长,并对PgAGO1蛋白的结构进行预测、氨基酸序列多重比对以及构建系统进化树等分析;同时采用实时定量PCR检测PgAGO1基因在人参不同组织根、茎、叶、花和愈伤中表达水平。结果:克隆的PgAGO1全长cDNA为3 776 bp,其中包括5'UTR 204 bp,3'UTR 254 bp,基因内部包含完整的开放阅读框,可编码1 105个氨基酸,预测蛋白质相对分子质量为122.22 kDa,理论等电点pI 9.71;实时定量PCR检测结果表明PgAGO1基因在人参花中的表达量最高,在根中最低。结论:从人参叶片中克隆得到PgAGO1的全长cDNA,为进一步研究该基因在人参组织发育中的调节作用打下基础。

Argonaute 1（AGO1） is a core component of the RNA-induced silencing complex（RISC） which plays a crucial role in small RNA-mediated gene silencing.AGO1 gene has been characterized in various plants,such as Arabidopsis and rice.However,there is no information about AGO1 in the medicinal plant species,Panax ginseng.Using the rapid amplification of cDNA ends technology（RACE）,we cloned full-length PgAGO1 cDNA from Panax ginseng.It is 3 776 bp in length,including 204 bp of 5′ UTR,254 bp of 3′ UTR,and 3 318 bp of ORF encoding 1 106 amino acids.The molecular weight（MW） and theroretical isoelectric point（pI） of the deduced PgAGO1 protein is 122.22 kDa and 9.71,respectively.PgAGO1 shares 91.72% similarity with Arabidopsis AtAGO1 and contains three consered domains,including DUF1785,PAZ and Piwi,suggesting it is an authentic AGO.PgAGO1 was expressed in all of the tissues analyzed with the highest level in flowers and the lowest level in roots.The results provide useful information for further elucidating the function of AGO1 in Panax ginseng.