【摘要】 干细胞治疗的关键是移植细胞可以在病变部位定植存活,通过多模态成像对移植细胞进行体内实时、定量的示踪,对于优化移植策略至关重要.其中,生物发光、PET成像和荧光成像是非常有效的多模态成像手段.为了实现多模态成像,我们采用自裂解多肽2A连接的方式,将3种成像技术对应的3个报告基因(Fluc、HSVttk、copGFP)同步表达.利用CRISPR/Cas9技术,以腺相关病毒整合位点1(Adeno-Associated Virus integration Site 1,AAVS1)为靶点,通过同源重组,在人胚胎干细胞H7中建立了不同启动子驱动多模态成像系统.我们发现CAG(Cytomegalovirus-immediate early enhancer/chickenβ-Actin/rabbitβ-Globin)启动子驱动的报告系统明显优于UB(Ubiquitin)启动子系统,且无沉默现象.更多还原

【Abstract】 The survival and retention are important for stem cell therapy.Multimodality imaging is powerful for tracking and quantifing the transplanted stem cells,providing a reliable way to optimize transplantaion strategy,including bioluminescence,PET imaging and Green Fluorescent Protein.In this study,Fluc,HSCttk and copGFP were constructed using 2 Apeptide fusion method for coexpression.By CRISPR/Cas9 technology,the three reporters drived by different promoters were targeted to AAVS1 site through homologous recombination.We found that the CAG(cytomegalovirus-immediate early enhancer/chickenβ-actin/rabbitβ-globin)promoter is more efficent to drive three reportors expression than Ubiquitin promoter,without gene silencing.更多还原