Dataset Information

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

ABSTRACT:

Background

The 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.

Results

LDH-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.

Conclusions

LDH-NSs have proven to be a highly effective molecular tool for delivering dsRNA into intact plant cells, enabling accurate control of target gene expression.

SUBMITTER: Zhang H

PROVIDER: S-EPMC10165820 | biostudies-literature | 2023 May

REPOSITORIES: biostudies-literature

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Publications

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

Zhang He H Li Xinyu X Yu Dong D Guan Junqi J Ding Hao H Wu Hongyang H Wang Qiang Q Wan Yinglang Y

Plant methods 20230508 1

<h4>Background</h4>The 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.<h4>Results</h4>LDH-NSs were synthesized by using the co-precipitation method, while the dsRNAs target ...[more]

PMID: 37158914

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Project description:BackgroundLayered double hydroxide lactate nanosheets (LDH-lactate-NS) are powerful carriers for delivering macro-molecules into intact plant cells. In the past few years, some studies have been carried out on DNA/RNA transformation and plant disease resistance, but little attention has been paid to these factors during LDH-lactate-NS synthesis and delamination, nor has their relationship to the DNA adsorption capacity or transformation efficiency of plant cells been considered.ResultsSince the temperature during delamination alters particle sizes and zeta potentials of LDH-lactate-NS products, we compared the LDH-lactate-NS stability, DNA adsorption rate and delivery efficiency of fluorescein isothiocyanate isomer I (FITC) of them, found that the LDH-lactate-NS obtained at 25 °C has the best characters for delivering biomolecules into plant cell. To understand the potential side effects and cytotoxicity of LDH-lactate-NS to plants, we compared the root growth rate between the Arabidopsis thaliana seedlings grown in the culture medium with 1-300 μg/mL LDH-lactate-NS and equivalent raw material, Mg(lactate)2 and Al (lactate)3. Phenotypic analysis showed LDH in a range of 1-300 μg/mL can enhance the root elongation, whereas the same concentration of raw materials dramatically inhibited root elongation, suggesting the nanocrystallization has a dramatical de-toxic effect to Mg(lactate)2 and Al (lactate)3. Since enhancing of root elongation by LDH is an unexpected phenomenon, we further designed experiments to investigate influence of LDH to Arabidopsis seedlings. We further used the gravitropic bending test, qRT-PCR analysis of auxin transport proteins, non-invasive micro-test technology and liquid chromatography-mass spectrometry to investigate the auxin transport and distribution in Arabidopsis root. Results indicated that LDH-lactate-NS affect root growth by increasing the polar auxin transport.ConclusionsOptimal synthesized LDH-lactate-NS can delivery biomolecules into intact plant cells with high efficiency and low cytotoxity. The working solution of LDH-lactate-NS can promote root elongation via increase the polar auxin transport in Arabidopsis roots.

Project description:Layered double hydroxides (LDHs) are highly effective drug delivery systems, owing to their capacity to intercalate or adsorb biomaterials, flexible structure, swelling property, high stability, good biocompatibility, and ease of synthesis. Phenytoin (PHT) is an antiseizure BCS (Biopharmaceutics Classification System) class II drug, presenting low aqueous solubility. Therefore, the current study aimed at increasing its solubility, dissolution, and bioavailability. PHT was intercalated to the MgAl-LDH formed in situ and successful intercalation to form MgAl-PHT-LDH was confirmed by FTIR, PXRD, DSC, and TGA. Examination of particle size and morphology (by photon correlation spectroscopy and electron microscopy, respectively) confirmed the formation and intercalation of nanostructured LDH. Intercalation enhanced the saturation solubility of PHT at 25°C in 0.1N HCl and phosphate buffer (pH 6.8) by 6.57 and 10.5 times respectively. The selected drug excipient powder blend for the formulation of MgAl-PHT-LDH tablets exhibited satisfactory properties in both pre-compression parameters (angle of repose, bulk density, tapped density, Carr's index, and Hausner ratio) and tablet characteristics (weight variation, thickness, hardness, friability, content uniformity, and disintegration time). MgAl-PHT-LDH tablets showed better dissolution of PHT compared to unprocessed PHT tablets at all time points. Oral bioavailability of MgAl-PHT-LDH tablets and unprocessed PHT tablets was tested in two groups of Sprague Dawley rats based on analysis of serum levels of both forms of PHT by UPLC-ESI-MS/MS serum. MgAl-PHT-LDH tablets demonstrated a relative bioavailability of 130.15% compared to unprocessed PHT tablets, confirming a significantly higher oral bioavailability of MgAl-PHT-LDH. In conclusion, MgAl-PHT-LDH could provide a strategy for enhancing solubility, dissolution, and thereby bioavailability of PHT, enabling the evaluation of theclinical efficacy of MgAl-PHT-LDH tablets for the treatment of seizures at lower PHT doses.

Dataset Information

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

Background

Results

Conclusions

Publications

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

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