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A vector-free gene interference system using delaminated Mg-Al-lactate layered double hydroxide nanosheets as molecular carriers to intact plant cells

文献类型： 外文期刊

作者： Zhang, He 1 ; Li, Xinyu 1 ; Yu, Dong 1 ; Guan, Junqi 1 ; Ding, Hao 1 ; Wu, Hongyang 3 ; Wang, Qiang 4 ; Wan, Yinglang 1 ;

作者机构： 1.Hainan Univ, Coll Trop Crops, Hainan Key Lab Sustainable Utilizat Trop Bioresour, Haikou 570228, Peoples R China

2.Chinese Acad Trop Agr Sci, Environm & Plant Protect Inst, Key Lab Integrated Pest Management Trop Crops, Minist Agr & Rural Affairs, Haikou 571101, Peoples R China

3.Chinese Acad Agr Sci, Inst Vegetables & Flowers, Beijing 100081, Peoples R China

4.Beijing Forestry Univ, Coll Environm Sci & Engn, 35 Qinghua East Rd, Beijing 100083, Peoples R China

关键词： RNA delivery; dsRNA; RNAi; Bioconjugates; Target gene expression

期刊名称：PLANT METHODS （ 影响因子：5.1； 五年影响因子：6.1 ）

ISSN：

年卷期： 2023 年 19 卷 1 期

页码：

收录情况： SCI

摘要： BackgroundThe Mg-Al-lactate layered double hydroxide nanosheet (LDH-NS) has shown great potential as an optimal nanocarrier for extensive use in plants. However, previous studies in plant sciences have not provided a clear description of the application for the LDH-NSs-based double-stranded RNA (dsRNA) delivery (LDH-dsRNA) system in different tissues of both model and non-model species.ResultsLDH-NSs were synthesized by using the co-precipitation method, while the dsRNAs targeting genes of interest were prepared in vitro using T7 RNA polymerase. The LDH-dsRNA bioconjugates with a neutral charge were produced by incubating with the mass ratio of LDH-NSs to dsRNA at 3:1, which were then introduced into intact plant cells using three different approaches, including injection, spray, and soak. The LDH-dsRNA delivery method was optimized by inhibiting the expression of the Arabidopsis thaliana ACTIN2 gene. As a result, soaking A. thaliana seedlings in a medium containing LDH-dsRNA for 30 min led to the silencing of 80% of the target genes. The stability and effectiveness of the LDH-dsRNA system were further confirmed by the high-efficiency knockdown of plant tissue-specific genes, including that encoding phytoene desaturase (PDS), WUSCHEL (WUS), WUSCHEL-related homeobox 5 (WOX5), and ROOT HAIR DEFECTIVE 6 (RHD6). In addition, the LDH-dsRNA system was employed in cassava, where it was found that the expression of the gene encoding nucleotide-binding site and leucine-rich repeat (NBS-LRR) was significantly reduced. As a result, the resistance of cassava leaves to pathogens was weakened. Noteworthy, the injection of LDH-dsRNA into leaves resulted in a significant downregulation of target genes in both stems and flowers, indicating the successful transport of LDH-dsRNA from leaves to other parts of plants.ConclusionsLDH-NSs have proven to be a highly effective molecular tool for delivering dsRNA into intact plant cells, enabling accurate control of target gene expression.

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