ShRNA-LDH纳米颗粒的制备及薇甘菊生物防治

Preparation of ShRNA-LDH nanoparticle and its application in biological control of Mikania micrantha

核糖核酸干扰(ribonucleic acid interference,RNAi)技术在植物保护中具有高效、安全和环保的特点.外源喷施短发夹RNA(short hairpin RNA,shRNA)可以有效沉默薇甘菊(Mikania micrantha)叶绿素a/叶绿素b结合蛋白相关基因.为提高喷施shRNA对薇甘菊的防治效果,制备直径为10~100 nm的层状双氢氧化物(layered double hydroxide,LDH)黏土纳米片为shRNA的载体,该载体具有良好的生物相容性,具环境友好.研究发现,LDH结合shRNA的最佳质量比为1∶10.shRNA喷施实验表明,有轻微创伤的薇甘菊叶片对shRNA更敏感,并且LDH直接喷施对薇甘菊的生长几乎没有任何影响.为比较LDH-shRNA和单独的shRNA对薇甘菊的控制效果,利用分别搭载了6个shRNA的LDH纳米颗粒或单独的shRNA喷施薇甘菊叶片,在喷施6 d后观察到叶片喷施LDH-shRNA有更加明显的坏死表型.研发的基于LDH的shRNA携带系统可显著提高RNAi在防控薇甘菊中的有效性.

Ribonucleic acid interference(RNAi)is a highly efficient,safe,and environmentally friendly method in plant protection.Exogenous spraying of short hairpin RNAs(shRNAs)targeting light-harvesting chlorophyll a or chlorophyll b binding protein genes onto the leaves of Mikania micrantha(M.micrantha)H.B.K.can efficiently silence the expression of these genes.To improve the effectiveness of shRNA-mediated control of M.micrantha,layered double hydroxide(LDH)clay nanosheets with a diameter between 10 to 100 nm and good biocompatibility and environmental friendliness were prepared as shRNA carriers in this study.We found that the optimal binding mass ratio of LDH to shRNA was 1∶10.ShRNA spraying assays showed that M.micrantha leaves with micro-wound were more sensitive to shRNA,and spraying LDH directly to M.micrantha had no effect on plant growth.To compare the effects of LDH-shRNA and shRNA alone on the control of M.micrantha,we sprayed LDH loaded with six different shRNA respectively or shRNA alone onto M.micrantha leaves,and a more severe necrotic phenotype was observed on the 6th day after LDH-shRNA-sprayed.Taken together,our study successfully developed an LDH-based shRNA carrying system to significantly improve the effectiveness of RNAi in the control of M.micrantha.