Preprint Article Version 1 Preserved in Portico This version is not peer-reviewed

Improved piggyBac Transformation with Capped Transposase mRNA in Pest Insects

Irina Haecker
* ORCID logo ,
Tanja Rehling
,
Henrik Schlosser
,
Daniela Mayorga-Churion
ORCID logo ,
Mara Heilig
ORCID logo ,
Ying Yan
ORCID logo ,
Peter Armbruster
ORCID logo ,
Marc F. Schetelig
ORCID logo
Version 1 : Received: 31 August 2023 / Approved: 31 August 2023 / Online: 1 September 2023 (14:00:54 CEST)

A peer-reviewed article of this Preprint also exists.

Häcker, I.; Rehling, T.; Schlosser, H.; Mayorga-Ch, D.; Heilig, M.; Yan, Y.; Armbruster, P.A.; Schetelig, M.F. Improved piggyBac Transformation with Capped Transposase mRNA in Pest Insects. Int. J. Mol. Sci. 2023, 24, 15155. Häcker, I.; Rehling, T.; Schlosser, H.; Mayorga-Ch, D.; Heilig, M.; Yan, Y.; Armbruster, P.A.; Schetelig, M.F. Improved piggyBac Transformation with Capped Transposase mRNA in Pest Insects. Int. J. Mol. Sci. 2023, 24, 15155.

Abstract

Creating transgenic insects is a key technology in insect genetics and molecular biology. A widely used instrument in insect transgenesis is the piggyBac transposase, resulting in essentially ran-dom genomic integrations. In contrast, site-specific recombinases allow the targeted integration of the transgene construct into a specific genomic target site. Both strategies, however, often face limitations due to low transgenesis rates. We aimed to enhance transgenesis rates by utilizing a capped mRNA as a source of transposase or recombinase instead of a helper plasmid. A system-atic comparison of transgenesis rates in Aedes mosquitoes, as models for hard to transform in-sects, showed that suppling piggyBac transposase as mRNA increased the average transfor-mation efficiency in Aedes aegypti from less than 5% with the plasmid source to about 50% with mRNA. Similar high transformation activity was observed in Ae. albopictus with pBac mRNA. No efficiency differences between plasmid and mRNA were observed in recombination experi-ments. Furthermore, a codon-optimized version of piggyBac transposase delivered as plasmid didn’t improve the transformation efficiency in Ae. aegypti or the agricultural pest D. suzukii. We believe that the use of mRNA has strong potential for enhancing pBac transformation efficiencies in other mosquitoes and important agricultural pests such as tephritids.

Keywords

Insect transgenesis; Aedes; Drosophila suzukii; tephritids; transformation efficiency; recombination efficiency; piggyBac transposase; helper plasmid; capped mRNA

Subject

Biology and Life Sciences, Insect Science

Copyright: This is an open access article distributed under the Creative Commons Attribution License which permits unrestricted use, distribution, and reproduction in any medium, provided the original work is properly cited.

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