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RecV 重组酶系统用于活体内单细胞或细胞群体的靶向光遗传学修饰。

RecV recombinase system for in vivo targeted optogenomic modifications of single cells or cell populations.

机构信息

Allen Institute for Brain Science, Seattle, WA, USA.

CNC Program, Stanford University, Palo Alto, CA, USA.

出版信息

Nat Methods. 2020 Apr;17(4):422-429. doi: 10.1038/s41592-020-0774-3. Epub 2020 Mar 23.

DOI:10.1038/s41592-020-0774-3
PMID:32203389
原文链接:https://pmc.ncbi.nlm.nih.gov/articles/PMC7135964/
Abstract

Brain circuits comprise vast numbers of interconnected neurons with diverse molecular, anatomical and physiological properties. To allow targeting of individual neurons for structural and functional studies, we created light-inducible site-specific DNA recombinases based on Cre, Dre and Flp (RecVs). RecVs can induce genomic modifications by one-photon or two-photon light induction in vivo. They can produce targeted, sparse and strong labeling of individual neurons by modifying multiple loci within mouse and zebrafish genomes. In combination with other genetic strategies, they allow intersectional targeting of different neuronal classes. In the mouse cortex they enable sparse labeling and whole-brain morphological reconstructions of individual neurons. Furthermore, these enzymes allow single-cell two-photon targeted genetic modifications and can be used in combination with functional optical indicators with minimal interference. In summary, RecVs enable spatiotemporally precise optogenomic modifications that can facilitate detailed single-cell analysis of neural circuits by linking genetic identity, morphology, connectivity and function.

摘要

脑回路由大量具有不同分子、解剖和生理特性的相互连接的神经元组成。为了允许针对单个神经元进行结构和功能研究，我们基于 Cre、Dre 和 Flp（RecVs）创建了光诱导的、特定于位点的 DNA 重组酶。RecVs 可以通过体内单光子或双光子光诱导来诱导基因组修饰。它们可以通过修饰小鼠和斑马鱼基因组中的多个基因座，对单个神经元进行靶向、稀疏和强烈的标记。与其他遗传策略相结合，它们允许不同神经元类别的交叉靶向。在小鼠皮层中，它们可以稀疏标记和重建单个神经元的全脑形态。此外，这些酶允许单细胞双光子靶向基因修饰，并且可以与具有最小干扰的功能性光学指示剂结合使用。总之，RecVs 实现了时空精确的光遗传学修饰，通过将遗传同一性、形态、连接和功能联系起来，有助于对神经回路进行详细的单细胞分析。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b3a6/7135964/459731eb4021/nihms-1560184-f0005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b3a6/7135964/df3aef3dd3bc/nihms-1560184-f0001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b3a6/7135964/267afef21088/nihms-1560184-f0002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b3a6/7135964/44f82fce8a8a/nihms-1560184-f0003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b3a6/7135964/a45864b37302/nihms-1560184-f0004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b3a6/7135964/459731eb4021/nihms-1560184-f0005.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b3a6/7135964/df3aef3dd3bc/nihms-1560184-f0001.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b3a6/7135964/267afef21088/nihms-1560184-f0002.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b3a6/7135964/44f82fce8a8a/nihms-1560184-f0003.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b3a6/7135964/a45864b37302/nihms-1560184-f0004.jpg
https://cdn.ncbi.nlm.nih.gov/pmc/blobs/b3a6/7135964/459731eb4021/nihms-1560184-f0005.jpg

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