Protocol for culturing and functionally manipulating planarian neoblasts using SiR-DNA-based flow cytometry
Wenya Zhang1, Xinran Li2, Yun Zhao3
1Westlake Laboratory of Life Sciences and Biomedicine, Key Laboratory of Growth Regulation and Translational Research of Zhejiang Province, School of Life Sciences, Westlake University, Hangzhou, Zhejiang 310024, China; Institute of Biology, Westlake Institute for Advanced Study, Hangzhou, Zhejiang 310024, China; School of Basic Medical Sciences, Fudan University, Shanghai 200030, China.
Abstract:
Neoblasts are the only cells capable of proliferation in planarians. The traditional flow cytometry protocol using Hoechst inhibits the cell cycle. Here, we present a protocol for culturing and functionally manipulating planarian neoblasts using SiR-DNA-based flow cytometry. We describe steps for cell dissociation and staining, flow cytometry, and cell collection and culture. We then detail procedures for Nanoluciferase mRNA transfection. This protocol facilitates further investigations into the pluripotency and regeneration mechanisms within neoblasts. For complete details on the use and execution of this protocol, please refer to Lei et al.1.
