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利用表达Cre重组酶的高容量腺病毒载体在体外和体内高效切除floxed序列。

Efficient in vitro and in vivo excision of floxed sequences with a high-capacity adenoviral vector expressing Cre recombinase.

作者信息

Badorf Michael, Edenhofer Frank, Dries Volker, Kochanek Stefan, Schiedner Gudrun

机构信息

Center for Molecular Medicine (ZMMK), University of Cologne, Cologne, Germany.

出版信息

Genesis. 2002 Jul;33(3):119-24. doi: 10.1002/gene.10099.

Abstract

Conditional gene expression or gene disruption using Cre/loxP- or FLP/frt-based recombination systems are valuable tools for studying gene function in development and disease. Recombinant adenoviral vectors expressing Cre recombinase have been suggested as an alternative for deletion of floxed sequences. To further improve this approach we generated a high-capacity adenoviral (HC-Ad) vector expressing Cre (HC-Adcre). In this vector all viral coding sequences are deleted resulting in decreased toxicity. In the present study HC-Adcre efficiently mediated recombination between two loxP sites located in the genome of a reporter cell line. When intravenously injected into ROSA26 reporter mice, a floxed sequence was excised in hepatocytes resulting in expression of the beta-gal reporter. Our data indicate that HC-Ad vectors expressing Cre effectively delete floxed sequences in vivo and have a significant potential as a tool for functional studies in mice.

摘要

利用基于Cre/loxP或FLP/frt重组系统的条件性基因表达或基因敲除是研究发育和疾病中基因功能的宝贵工具。表达Cre重组酶的重组腺病毒载体已被提议作为删除floxed序列的替代方法。为了进一步改进这种方法,我们构建了一种表达Cre的高容量腺病毒(HC-Ad)载体(HC-Adcre)。在该载体中,所有病毒编码序列均被删除,从而降低了毒性。在本研究中,HC-Adcre有效地介导了位于报告细胞系基因组中的两个loxP位点之间的重组。当静脉注射到ROSA26报告小鼠体内时,肝细胞中的一个floxed序列被切除,导致β-半乳糖苷酶报告基因表达。我们的数据表明,表达Cre的HC-Ad载体在体内能有效删除floxed序列,作为小鼠功能研究工具具有巨大潜力。

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