Kim, Seung-Chan Kim Ajung Park, Jae-Yong Hwang, Eun Mi 2024-01-19T13:00:20Z 2024-01-19T13:00:20Z 2022-02-17 2022-02 0165-0270 https://pubs.kist.re.kr/handle/201004/115771 Background: RNA interference (RNAi) is a powerful technique to effectively silence or knock down gene function in mammalian cells. For better cell-type RNAi experiments in vivo, AAV vector-based RNA interference systems need to be improved. New method: In this study, we developed an AAV vector (CREon shRNA) that expressed CRE-dependent short hairpin RNA (shRNA) and fluorescent proteins simultaneously. Results: We verified the Cre-dependent knockdown efficiency of the newly developed CREon shRNA vector in both HEK293T cells overexpressing TREK-1 and PC3 cells with endogenous TREK-1. Next, we packaged this TREK-1 CREon vector with AAV and injected it into the hippocampus of the brain together with a synapsin or GFAP promoter-driven CRE virus, confirming that it works well cell-selectively even in vivo. Finally, this viral vector was applied to an animal model of LPS-induced depression to determine whether behavioral changes occurred. Comparison with existing methods: With the existing pSico or pAAV-Sico-Red vectors, expression of fluorescent protein disappears when shRNA is conditionally activated by CRE recombinase, but our Creon shRNA vector showed simultaneous expression of both shRNA and fluorescent protein. Thus, it offers the advantage of allowing easy visual distinction of knocked-down cells. Conclusion: The newly improved CREon shRNA vector can be used as a novel research tool for conditional shRNA, and may be useful for various in vivo studies such as cancer and neurobiology. ？ 2021 Elsevier B.V. English Elsevier BV Improved AAV vector system for cell-type-specific RNA interference Article 10.1016/j.jneumeth.2021.109452 1 Journal of Neuroscience Methods, v.368 Journal of Neuroscience Methods 368 N scie scopus 000784382400001 2-s2.0-85121974964 Biochemical Research Methods Neurosciences Biochemistry & Molecular Biology Neurosciences & Neurology Article POL-II PROMOTER EXPRESSION CHANNEL TARGET TREK-1 SHRNA AAV vector Cell-type specific Conditional shRNA TREK-1