# PiggyBac-mediated transgenesis and CRISPR–Cas9 knockout in the greater wax moth, Galleria mellonella

## TL;DR

Researchers developed genetic tools for the greater wax moth, enabling transgenesis and gene editing to enhance its use in infection research.

## Contribution

A robust method for PiggyBac-mediated transgenesis and CRISPR–Cas9 gene knockout in Galleria mellonella is introduced.

## Key findings

- Transgenic G. mellonella lines were successfully generated using the PiggyBac transposon system.
- CRISPR–Cas9 technology enabled precise gene knockouts in the species.
- These genetic tools expand the utility of G. mellonella as a model for infection studies.

## Abstract

The larvae of the greater wax moth, Galleria mellonella, are gaining prominence as a versatile nonmammalian in vivo model to study host–pathogen interactions. Their ability to be maintained at 37 °C, coupled with a broad susceptibility to human pathogens and a distinct melanization response that serves as a visual indicator for larval health, positions G. mellonella as a powerful resource for infection research. Despite these advantages, the lack of genetic tools, such as those available for zebrafish and Drosophila melanogaster, has hindered development of the full potential of G. mellonella as a model organism. Here we describe a robust methodology for generating transgenic G. mellonella using the PiggyBac transposon system and for precise gene knockouts via CRISPR–Cas9 technology. These advances significantly enhance the utility of G. mellonella in molecular research, paving the way for its widespread use as an inexpensive and ethically compatible animal model in infection biology and beyond.

The authors used the PiggyBac and CRISPR–Cas9 systems to generate transgenic lines and gene knockouts in the greater wax moth, Galleria mellonella. These approaches are expected to broaden the range of applications for this model in preclinical research.