RecV recombinase system for in vivo targeted optogenomic modifications of single cells or cell populations
Yao, S.; Yuan, P.; Ouellette, B.; Zhou, T.; Mortrud, M.; Balaram, P.; Chatterjee, S.; Wang, Y.; Daigle, T. L.; Tasic, B.; Kuang, X.; Gong, H.; Luo, Q.; Zeng, S.; Curtright, A.; Dhaka, A.; Kahan, A.; Gradinaru, V.; Chrapkiewicz, R.; Schnitzer, M.; Zeng, H.; Cetin, A.
Show abstract
Brain circuits are composed of vast numbers of intricately interconnected neurons with diverse molecular, anatomical and physiological properties. To allow highly specific targeting of individual neurons for structural and functional studies, we modified three site-specific DNA recombinases, Cre, Dre and Flp, by combining them with a fungal light-inducible protein, Vivid, so that their recombinase activities can be driven by blue light. We generated viral vectors to express these light-inducible recombinases and demonstrated that they can induce genomic modifications in dense or sparse populations of neurons in live mouse brains controlled by one-photon or two-photon light induction. As an important application, we showed that light-inducible recombinases can produce highly targeted, sparse and strong labeling of individual neurons thereby enabling whole-brain morphological reconstruction to identify their axonal projection specificity. In addition to targeting cortical brain areas, we applied the method in deep targets, with a demonstration of functional calcium imaging. These molecular tools enable spatiotemporally-precise, targeted genomic modifications that will greatly facilitate detailed analysis of neural circuits and linking genetic identity, morphology, connectivity and function.
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