EDGE CT: Developing transgenic and lineage tracing tools in planarians

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Planarians have captured the imagination of generations of scientists, and today play a critical role in our efforts to understand development and regeneration. These small flatworms can regenerate their entire body from small tissue fragments. As a result of the current surge in the sequencing technology, tremendous progress has been made over the past two decades to understand planarian biology from a variety of perspectives, including but not limited to regeneration, stem cell, animal body plan and evolution, germline, nervous system, metabolism, innate immunity, signaling pathways, regulatory RNAs, and genome stability. Yet the entire field has been limited by the lack of tools for transgene expression and genome editing. At the same time, technological innovation has exploded for transfection and gene editing of mammalian cells. In particular, nanotechnological approaches can very locally disrupt the cell membrane and inject molecular cargo directly, providing both low cell toxicity and high delivery efficiencies. In this project, the researchers will use the novel nanotechnology, "nanostraws," to deliver genetic materials into planarian cells, and provide a simple and effective means for transgene expression and genome editing. This project also aims to rapidly disseminate the new technology by facilitating data, protocol, and materials sharing across labs, and organizing training workshops. In addition, because of planarians' appealing regenerative ability, childlike cuteness, and ease to rear in the lab, the researchers will promote the usage of planarians in teaching, education, and outreach activities to engage non-scientists' interests in modern biological research.Planarians have been a powerful animal model to study tissue regeneration and stem cell biology. They have the unique capacity to regenerate the entire body from minute tissue fragments using pluripotent somatic stem cells. During this process, they reset the body axes and rebuild all organs in appropriate proportions. Although extensive genomic and transcriptomic information is available, progress on addressing causal genotype-phenotype relationships in planarians has been severely hindered by a lack of transgenic tools despite decades of attempts. Enabled by the recent technical advances made in the investigators' laboratories, including cell culture and transplantation methods, novel nanotechnology for gene delivery in hard-to-transfect cell types, and successful expression of luminescence reporters in planarian cells, they aim to address this long-standing challenge. This research project aims to develop and disseminate the methodology and resources for planarian transgenesis. Specifically, the goals of this project include (1) to develop nanoscale electroporation methods for transgene delivery in planarian cells, (2) to demonstrate genome editing using CRISPR/Cas9, and (3) to enhance the collaboration, training, and diversity of the planarian research community to address a broad spectrum of questions in molecular, cellular, organismal, and evolutionary biology.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.