TY  - JOUR
TI  - Introducing Exogenous DNA Vectors Directly into Trypoxylus dichotomus Larvae Via In Vivo Electroporation
AU  - Morita, Shinichi 
AU  - Niimi, Teruyuki 
VL  - 16
IS  - 4
PY  - 2026
DA  - 2026/02/20
SP  - e5602
C1  - Bio-protocol 2026;16:e5602
DO  - 10.21769/BioProtoc.5602
UR  - https://doi.org/10.21769/BioProtoc.5602
AB  - In the Japanese rhinoceros beetle Trypoxylus dichotomus, gene function studies have relied mainly on systemic larval RNA interference (RNAi), as gain-of-function techniques remain underdeveloped and germline transgenesis is impractical given the species’ approximately one-year generation time. In addition, because larval RNAi is systemic, it has been difficult to analyze the function of lethal genes. Here, we present a simple and efficient protocol for the direct introduction of exogenous DNA into T. dichotomus larvae via in vivo electroporation. This protocol includes optimized procedures for adult breeding and egg collection, as well as a rigorously parameterized electroporation technique that delivers a piggyBac transposon vector into region-specific larval tissues. Within one day after electroporation, treated larvae exhibit mosaic expression of a reporter gene, enabling rapid tissue-specific functional analysis without the need to establish stable germline transgenic lines. Moreover, the key promoter used in this system (T. dichotomus actinA3 promoter) is effective across diverse insect species, indicating that the method can be readily adapted to other non-model insects. Overall, this electroporation-based approach provides a valuable gain-of-function tool for T. dichotomus and potentially many other insect species.
KW  - Electroporation
KW  - Japanese rhinoceros beetle
KW  - Trypoxylus dichotomus
KW  - piggyBac
JF  - Bio-protocol
SN  - 2331-8325
PB  - Bio-protocol LLC.
BIO101  - False