Sheet-like clay nanoparticles deliver RNA into developing pollen to efficiently silence a target gene

Area of Science:

• Plant biotechnology
• Nanotechnology
• Molecular biology

Background:

• RNA interference (RNAi) offers a method for modifying plant traits without genetic modification.
• Efficient delivery of double-stranded RNA (dsRNA) into plant cells is a significant challenge for RNAi applications.
• Developing plant cells, like tomato pollen, serve as valuable models for studying cellular uptake mechanisms.

Purpose of the Study:

• To investigate the efficacy of layered double hydroxide (LDH) nanoparticles as a nonviral vector for dsRNA delivery into plant cells.
• To evaluate the potential of LDH nanoparticles for inducing RNA interference (RNAi) in tomato pollen.
• To determine the optimal conditions for LDH-mediated dsRNA delivery and gene silencing.

Main Methods:

• Utilized developing tomato (Solanum lycopersicum) pollen as a model system for cellular uptake studies.
• Employed layered double hydroxide (LDH) nanoparticles (up to 50 nm) for the delivery of dsRNA.
• Quantified transgene reporter mRNA levels using quantitative PCR to assess gene silencing efficiency.

Main Results:

• LDH nanoparticles were readily internalized by early bicellular tomato pollen through energy-dependent and -independent pathways.
• Efficient dsRNA delivery into tomato pollen was achieved within 2-4 hours of incubation with LDH nanoparticles.
• An 89% decrease in transgene reporter mRNA was observed in early bicellular pollen, significantly higher than the 37% reduction with naked dsRNA.
• Gene silencing efficacy was found to be dependent on LDH particle size, dsRNA dose, LDH-dsRNA complexing ratio, and treatment duration.

Conclusions:

• LDH nanoparticles represent a highly effective nonviral vector system for delivering dsRNA into plant cells.
• This method facilitates efficient RNA interference in plant systems without the need for genetic modification.
• LDH nanoparticles hold promise for the targeted delivery of various biomolecules in plant biotechnology applications.