内含肽介导的Split-Cre系统的构建

Construction of an intein-mediated Split-Cre system

摘要:

Split-Cre系统是指被拆分为无活性的N端和C端这2个片段的Cre酶在一定条件下重组为有酶活性的全长Cre的基因工程工具。该系统常结合LoxP元件使用，近年来在多基因操纵、多基因活性标记以及神经环路示踪等方面有着重要的应用。为了开发更高效的Split-Cre系统，本研究利用海洋红嗜热盐菌(Rhodothermus marinus,Rma)内含肽对Cre进行拆分，在“红绿灯”报告细胞系中筛选出了能够高效介导Cre酶重组的S102拆分位点，并且通过双腺相关病毒(adeno-associated virus,AAV)将S102 Split-Cre系统递送至小鼠体内，证明了Rma内含肽介导的S102 Split-Cre系统在小鼠体内同样具有高效的活性。本研究为Split-Cre的基础和应用研究提供了可靠的参考。

Abstract:

The Split-Cre system consists of two inactive polypeptides: NCre and CCre, which can be recombined into an active full-length Cre under certain conditions. This system is typically used with LoxP. To develop an efficient Split-Cre system, this study used Rma intein from Rhodothermus marinus to split Cre and screened out the split site S102 which could efficiently mediate the recombination of Cre in the “Traffic Light” reporter cell line. Moreover, the S102 Split-Cre system was delivered to mice by dual-adeno-associated virus (AAV), and it was demonstrated that the efficiency of the Rma intein-mediated S102 Split-Cre system was comparable to the full-length Cre in mice. This system lays a foundation for both basic and applied research on Split-Cre.