Establishment of a tTA-dependent photoactiveatable Cre recombinease knock-in mouse model for optogenetic genome engineering.
Establishment of a tTA-dependent photoactiveatable Cre recombinease knock-in mouse model for optogenetic genome engineering.
复制标题
建立用于光遗传学基因组工程的 tTA 依赖性光活化 Cre 重组酶敲入小鼠模型。
DOI：
10.1016/j.bbrc.2020.03.015
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发表时间：
2020
期刊：
影响因子：
--
通讯作者：
T.
中科院分区：
文献类型：
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作者：
9.Takao;T.;Hiraoka;Y.;Kawabe;K.;Yamada;D.;Ming;L.;Tanaka;K.;Sato;M.;Takarada;T.
Cre-loxP重组系统广泛用于产生用于生物医学研究的遗传修饰小鼠。最近，一个高效的光活化的Cre（PA-Cre）的基础上重新组装的分裂Cre片段已经建立。该技术能够实现高效的DNA重组，其在蓝光照射时以时空精度被激活。在这项研究中，我们使用CRISPR/Cas9系统产生了tTA依赖性可光活化的Cre-loxP重组酶敲入小鼠模型（TRE-PA-Cre小鼠）。将这些小鼠与ROSA 26-tdTomato小鼠（Cre报告小鼠）杂交，以观察由tdTomato表达标记的DNA重组。我们证明了外部非侵入性LED蓝光照明允许使用流体动力学尾静脉注射在用tTA表达载体转染的TRE-PA-Cre：ROSA 26-tdTomato小鼠的肝脏中进行有效的DNA重组。本研究建立的TRE-PA-Cre小鼠有望在体内以非侵入性、时空特异性和细胞类型特异性的方式进行光遗传基因组工程。
The Cre-loxP recombination system is widely used to generate genetically modified mice for biomedical research. Recently, a highly efficient photoactivatable Cre (PA-Cre) based on reassembly of split Cre fragments has been established. This technology enables efficient DNA recombination that is activated upon blue light illumination with spatiotemporal precision. In this study, we generated a tTA-dependent photoactivatable Cre-loxP recombinase knock-in mouse model (TRE-PA-Cre mice) using a CRISPR/Cas9 system. These mice were crossed with ROSA26-tdTomato mice (Cre reporter mouse) to visualize DNA recombination as marked by tdTomato expression. We demonstrated that external noninvasive LED blue light illumination allows efficient DNA recombination in the liver of TRE-PA-Cre:ROSA26-tdTomato mice transfected with tTA expression vectors using hydrodynamic tail vein injection. The TRE-PA-Cre mouse established here promises to be useful for optogenetic genome engineering in a noninvasive, spatiotemporal, and cell-type specific mannerin vivo.