Enhanced dsRNA Production via a Three-Terminator Vector and Transcriptomic Correlates of RNAi Exposure in Thrips

Area of Science:

• Entomology
• Molecular Biology
• Pest Management

Background:

• RNA interference (RNAi) shows potential for insect pest control.
• Efficient double-stranded RNA (dsRNA) uptake is crucial for RNAi efficacy but poorly understood in thrips.
• Optimized dsRNA production is necessary for functional studies.

Purpose of the Study:

• To enhance dsRNA production using an optimized E. coli expression system.
• To identify a conserved muscle actin fragment for dsRNA synthesis.
• To evaluate RNAi efficacy and dsRNA uptake mechanisms in thrips species (Megalurothrips usitatus and Frankliniella occidentalis).

Main Methods:

• Optimized E. coli expression system with a three-tandem terminator vector for increased dsRNA yield.
• Synthesized dsRNA targeting a conserved muscle actin gene.
• Administered dsRNA orally to thrips and assessed mortality, gene expression (RT-qPCR), and transcriptomic changes (RNA-seq).

Main Results:

• dsRNA yield increased approximately 11-fold with the optimized E. coli system.
• dsRNA targeting muscle actin caused concentration-dependent mortality (72% in M. usitatus, 48% in F. occidentalis) and significant mRNA down-regulation within 72 hours.
• Transcriptomic analysis suggested clathrin-mediated endocytosis and SID-1-like transport as potential dsRNA uptake pathways.

Conclusions:

• The optimized E. coli system facilitates efficient dsRNA production for RNAi studies in thrips.
• Muscle actin dsRNA effectively suppresses gene expression and causes mortality in target thrips species.
• Findings suggest a potential dual-pathway model for dsRNA uptake in thrips, informing future RNAi-based pest control strategies.