期刊
MOLECULAR REPRODUCTION AND DEVELOPMENT
卷 72, 期 1, 页码 54-67出版社
WILEY-LISS
DOI: 10.1002/mrd.20323
关键词
Cre-/IoxP system; conditional; diphtheria toxin; EGFP; genetic ablation; transgenic
Diphtheria, toxin A-chain (DT-A) is a potent inhibitor of protein synthesis. As little as a single molecule of DT-A can result in cell death. DT-A gene driven by a tissue-specific promoter is used to achieve genetic ablation of a particular cell lineage. However, this transgenic approach often results in aberrant depletion of unrelated cells. To avoid this, we established a method for specific depletion of a cell population by controlled expression of the DT-A gene via the Cre-IoxP system. We produced five transgenic mice carrying CETD construct containing IoxP-flanked enhanced green fluorescent protein (EGFP) cDNA and the DT-A gene. Transfection of primary cultured cells derived from CETD transgenic fetus with Cre expression plasmid resulted in extensive cell loss, as expected. Bigenic (double transgenic) offspring obtained by crossbreeding between CETD and MNCE transgenic mice in which Cre expression is controlled by the myelin basic protein (MBP) promoter exhibited embryonic lethality, suggesting expression of Cre at embryonic stages. Intravenous injection of Cre expression vector to CETD mice led to generation of glomerular lesions, probably due to predominant depletion of glomerular epithelial cells. This Cre-IoxP-based cell ablation technology is powerful and convenient method of generating mice lacking any chosen cell population.
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