# Assessment of Ubiquitous Promoters Driving Fluorescent Marker and Transposase Expression to Develop a High-Performance piggyBac Transgenic System in Bactrocera dorsalis

## TL;DR

Researchers developed a high-efficiency transgenic system in oriental fruit flies using strong promoters to drive fluorescent marker expression, enabling better genetic control methods.

## Contribution

The study identifies and utilizes the 3.6-kb BdPUb promoter for a high-performance piggyBac transgenic system in Bactrocera dorsalis.

## Key findings

- The 5.0-kb BdActA3a and 3.6-kb BdPUb promoters showed significantly stronger activity than their truncated variants.
- A fluorescent transgenic strain was developed with a transformation efficiency of approximately 26%.
- The transgenic strain exhibited stable inheritance and no negative impact on fecundity.

## Abstract

This study aimed to identify high-efficiency promoters in Bactrocera dorsalis. We compared the ability of different Actin and polyubiquitin-C (PUb) promoters to drive the expression of the fluorescent protein mScarlet-I in B. dorsalis embryos. The results showed that the 5.0-kb BdActA3a and 3.6-kb BdPUb promoters had significantly stronger activity than their truncated variants, and the BdPUb promoter showed particularly high performance. Using the 3.6-kb BdPUb promoter and piggyBac-mediated transgenesis technology, we successfully developed a high-performance piggyBac-mediated transgenic system and constructed a fluorescent transgenic strain with a transformation efficiency of approximately 26%. This strain displayed stage-specific fluorescence expression, stable inheritance, and no negative impact on fecundity. Our findings lay a solid foundation for developing genetically modified strains for the genetic control of B. dorsalis in the future.

Bactrocera dorsalis (oriental fruit fly) is a destructive invasive pest threatening global agriculture. Although integrated pest management is applied, environmentally friendly genetic control methods are urgently needed. The development of such methods particularly relies on efficient genetic elements. In this study, we compared the transient expression of mScarlet-I driven by various Actin and PUb promoters in B. dorsalis embryos. The truncation of two strong promoters, BdActA3a and BdPUb, revealed that the 5.0-kb BdActA3a and 3.6-kb BdPUb promoters drove significantly higher expression than their truncated variants. Notably, the BdPUb promoter was highly effective in driving fluorescent protein expression in B. dorsalis. Using the 3.6-kb BdPUb promoter, we constructed a transposase plasmid BdPUb-3.6 kb>hyPBase. By co-injecting the BdPUb 3.6kb>mScarlet-I donor construct, we successfully generated a fluorescent transgenic strain with a transgenic efficiency of approximately 26%. The strain exhibited stage-specific fluorescence and maternal effect and the homozygotes showed fecundity comparable to wild-type controls. The high performance of the piggyBac transposase and the fluorescence screening system provides a substantial technical foundation for basic research and future development of genetically modified strains to control B. dorsalis.