High resolution atomic force microscopy of double-stranded RNA.

Area of Science:

• Molecular Biology
• Biophysics

Background:

• Double-stranded RNA (dsRNA) plays critical roles in gene regulation, viral replication, and innate immunity.
• Its mechanical properties and structure are of significant interest for biotechnological applications.
• Previous studies have achieved high-resolution imaging of double-stranded DNA (dsDNA) using Atomic Force Microscopy (AFM), but dsRNA remains less characterized.

Purpose of the Study:

• To visualize the double helix structure of dsRNA under near-physiological conditions.
• To achieve sufficient resolution to identify the A-form sub-helical pitch periodicity.
• To identify the limiting factors for high-resolution AFM imaging of soft materials in liquid.

Main Methods:

• Utilized Atomic Force Microscopy (AFM) with high-sensitive force-detection methods.
• Performed imaging of dsRNA in a liquid medium under near-physiological conditions.
• Employed different imaging modes to assess their impact on resolution.

Main Results:

• Successfully visualized the dsRNA double helix at high resolution.
• Resolved the A-form sub-helical pitch periodicity of dsRNA.
• Demonstrated that tip-sample force and tip apex sharpness are the primary limitations for high-resolution AFM imaging in liquid, rather than the imaging mode itself.

Conclusions:

• High-resolution AFM imaging of dsRNA is achievable under near-physiological conditions.
• The study identifies critical parameters (force and tip sharpness) essential for optimizing AFM imaging of soft biological molecules.
• These findings pave the way for more detailed structural and mechanical characterization of dsRNA and similar biomolecules.