敲除和过表达miRNA446对其靶基因APP1的表达模式影响

Overexpression and inhibition of miR446 to study the expression pattern of its target gene APP1

在本课题组前期研究中,通过基因芯片数据发现了一系列在胁迫应答中表达量有变化的microRNA.其中以miRNA446的变化尤为显著.为进一步探求miRNA446在胁迫应答中的变化与功能,本实验通过在水稻中分别敲除和过表达miR446,检测干旱和冷胁迫下,miR446及其靶基因APP1的表达模式.结果表明,12h内野生型株系miR446的表达量升高至未胁迫的3至5倍,过表达株系中则是先降至低于未受胁迫时的50%后逐步上升,而APP1基因由于受到高水平miR446的剪切作用而一直降低,12h可降低至未受胁迫时的50%以下.敲除株系中,miR446的表达量被完全抑制,在整个胁迫过程中均保持接近于零的水平,而APP1在胁迫中也仅有微弱的先降后升趋势.

MicroRNA is a kind of small non-coding RNA, involved in plant growth and development, stress response and other physiological processes. In our previous study, a series of microRNA had been found that changed expression in the stress response on the basis of gene microarray. Among them, miR 446 is particularly significant. To further explore the change and function of miRNA446 in the stress re- sponse, the expression pattern of its target gene APP1 under drought and cold stress had been studied by overexpression and inhibition of miR446 in rice. The results showed that in the overexpression rice, the expression of miR446 increased after the first drop while in wild-type miR446 incrased 3 to 5 times higher in 12h,and APP1 gene decreased more than 50% in 12h due to the splicing action of high levels miR446. In the knockout rice, the expression of miR446 was completely inhibited in the whole process, remained close to zero, and APP1 increased after the first drop in a weak trend.