Development of a Transient HEK293 Expression System for Functional Analysis of Insect Odorant Receptors

Diplôme Master Development of a Transient HEK293 Expression System for Functional Analysis of Insect Odorant Receptors eng 2025-09-05 2025-09 author Garibian Armen 901292 director Rodriguez Ivan 28183 Département de génétique et évolution (Genev) (BIANI) 24 Mammalian cell systems provide a defined and scalable environment for functional analysis of insect odorant receptors (ORs), but many ORs remain unresponsive when expressed in this context. Stable HEK293 lines have been used successfully, yet they are time-consuming to generate and costly to maintain. To address these limitations, we developed a transient expression system in HEK293 cells and tested whether combining multiple construct-level optimizations reported in the literature could improve functional expression. The design integrates codon optimization, chimeric introns, N-terminal trafficking tags, a bidirectional promoter for OR and Orco co-expression, and a GCaMP6s indicator fused to Orco. Transfection and imaging protocols were refined in 96-well format to improve viability and reproducibility. Functional responses were robustly observed for OR47a, in line with published in vivo and in vitro data, whereas other tested receptors did not yield detectable activation under these conditions. These results demonstrate that combining established design strategies improves expression outcomes but does not universally restore receptor function, highlighting that some ORs may depend on additional cellular factors absent from mammalian systems. Dewey 590 MGBDE 44 Département de génétique et évolution 162 false 820 Groupe Rodriguez f0c8695c-19b8-4a9f-bfd4-40668580a34a Master thesis Restricted Master_Armen_Garibian.pdf 3131999 application/pdf 1fa7a06bbdf58a4a3b25c81ead685ddefe46c442a4aae2e4cada536871bb9e70 restrictedAccess Faculté des sciences / Section de biologie / Département de génétique et évolution

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