D3042: Exploring vivo-morpholino knockdown of piRNA associated genes Argonaute3 and Zucchini as an alternative to traditional dsRNAi in Aedes aegypti

RNA interference (RNAi) is a useful method for studying gene function, typically involving the introduction of double-stranded RNA (dsRNA) into cells or organisms. However, in organisms like Aedes aegypti mosquitoes, which have tight junctions in their germline tissues, delivering dsRNA with successful knockdown is challenging. Here, we explore vivo morpholino knockdown as an alternative to injecting dsRNA for targeting piRNA-associated genes for Argonaute3 and Zucchini in adult female Aedes aegypti. Using vivo morpholinos, which function in both the nucleus and cytosol, we aim to overcome the challenge of dsRNA penetration into germline tissues. We designed specific vivo morpholino molecules targeting Ago3 and Zuc, genes that play roles in the piRNA pathway and increase in expression 48 hours after a blood meal. These molecules are delivered via injection into adult mosquitoes. We hypothesize that vivo morpholinos may show better effectiveness in carcass tissues due to their broader cellular activity. Moreover, we are investigating whether vivo morpholinos can penetrate germline tissues effectively, considering factors like stability and limited immune response compared to dsRNA. Using quantitative PCR (qPCR), we compare the efficacy of vivo morpholino knockdown in carcass and ovary tissues to that of dsRNA injections. This study sheds light on the potential of vivo morpholinos as a viable alternative to dsRNA for gene knockdown studies in organisms that exhibit high levels of RNase activity like Aedes aegypti.