Improved piggyBac Transformation with Capped Transposase mRNA in Pest Insects

Area of Science:

• Insect genetics and molecular biology
• Transgenic insect technology

Background:

• Transgenic insect creation is vital for genetic research.
• PiggyBac transposase and site-specific recombinases are common tools, but often yield low efficiencies.
• Current methods using helper plasmids can be inefficient for hard-to-transform insects.

Purpose of the Study:

• To enhance insect transgenesis efficiencies.
• To evaluate the use of capped mRNA as an alternative to helper plasmids for delivering transposase or recombinase.

Main Methods:

• Systematic comparison of transgenesis efficiencies in Aedes mosquitoes (Aedes aegypti and Ae. albopictus).
• Delivery of piggyBac transposase and recombinase via capped mRNA versus helper plasmid.
• Testing a hyperactive piggyBac transposase variant delivered as a plasmid in Aedes aegypti and Drosophila suzukii.

Main Results:

• Supplying piggyBac transposase as mRNA increased transformation efficiency in Aedes aegypti from <5% to ~50%.
• Similar high efficiency was observed in Ae. albopictus using piggyBac mRNA.
• No efficiency difference was found between mRNA and plasmid for recombinase experiments; hyperactive piggyBac plasmid did not improve efficiency.

Conclusions:

• Messenger RNA (mRNA) delivery significantly enhances piggyBac transposase-mediated transgenesis efficiency in mosquitoes.
• This mRNA approach holds strong potential for improving transgenesis in other mosquito species and agricultural pests like tephritids.
• The study highlights mRNA as a superior source for piggyBac transposase delivery compared to traditional plasmid methods.