miR158调节油菜作物响应非生物胁迫作用机理的研究

miRNAs are a distinct class of small RNAs in plants that play an important role inthe regulation of plant environment stresses. Recently, we identified miR158 and its target DEAD-box ATP-dependent RNA helicase 6, from Brassica napus. Our previous experiments have demonstrated that miR158 was up-regulated in expression under Cd, NaCl and Cold stresses in Brassica napus, and its targets BnaA04g26450D and BnaC04g50480D was down-regulated in expression under these abiotic stresses. In addition, it was reported RNA helicases play an important role in abiotic stresses. In this study, we will construct overexpression vectors of the miR158 and and its target and transform them into Brassica napus, respectively. The expression patterns of miR158 and its target, the characteristics of phenotype, biomass and mechanisms of physiology and biochemistry will be studied under heavy metal Cd stress in the control and transgenic plants. The study is based on the molecular mechanism of miR158 induced by cadmium. The results also may provide the evidences in the role of miR158 and its target under Cd stress in Brassica napus.

miRNA是一类植物中独特的小分子RNA，它在调控植物逆境胁迫中发挥重要作用。最近，申请人通过高通量测序鉴定了油菜中的miR158及其靶基因——RNA解旋酶(DEAD-box ATP-dependent RNA helicase 6 )。初步研究表明，油菜miR158受镉，盐和冷害等非生物胁迫的诱导表达，而其靶基因BnaA04g26450D和BnaC04g50480D在镉等非生物胁迫下表达下调，并且相关报道表明RNA解旋酶在非生物胁迫中发挥重要作用。本项目拟通过构建miR158和及其靶基因的过表达载体，转化获得油菜转基因植株，探索miR158及其靶基因在镉等非生物胁迫下的表达模式、转基因植株的表型变化、生物量的改变及其在重金属镉等非生物胁迫下的生理生化反应。本项目立足于研究油菜miR158受镉等非生物胁迫的分子机理，同时将进一步探索油菜中miR158及其靶基因在非生物胁迫下所发挥的作用。

microRNA (miRNA) 是一类植物中独特的小分子RNA，它在调控植物逆境胁迫中发挥重要作用。本项目鉴定了油菜miR158及其靶基因在非生物胁迫下的功能。首先，我们通过降解组测序发现油菜BnRH24基因可被miR158剪切，是miR158的靶基因之一。进一步的表达分析发现miR158可被镉胁迫诱导，而其靶基因BnRH24的表达受到镉胁迫的抑制。通过高通量测序发现，pre-miR158a的3’末端存在尿苷化和胞苷化修饰，并引发miR158/pre-miR158的降解，从而影响miR158的表达丰度。另外，我们对各转基因植株在ABA处理下的表型变化进行分析，发现35S:miR158a/b对ABA敏感，而STTM158对ABA不敏感，说明miR158可能通过ABA途径参与到非生物胁迫中。进一步分析miR158在非生物胁迫下的反应，发现在拟南芥和油菜中35S:miR158a/b对盐胁迫的耐受性下降，而STTM158对盐胁迫的耐受性上升。同时，我们发现在镉胁迫下，35S: BnRH24植株根长变短，叶片中Chl-a和Chl-b的浓度下降，说明BnRH24表达能够弱化植物对镉胁迫的耐受性。通过NBT和DAB染色实验，发现BnRH24促使植物组织中H2O2的增加，表明BnRH24的表达通过增加O2-的产生而有助于胁迫应激反应。综上所述，在本课题中，我们鉴定了miR158及其靶基因在非生物胁迫下的表达模式，并证明了miR158可调节其靶基因BnRH24改变植物对盐和镉等非生物胁迫的耐受能力。