Enhanced purification of cell-permeant Cre and germline transmission after transduction into mouse embryonic stem cells

  • Continuous expression of Cre recombinase has the potential to yield toxic side effects in various cell types, thereby limiting applications of the Cre/loxP system for conditional mutagenesis. In this study, we investigate the potential of Cre protein transduction to overcome this limitation. COS-7, CV1-5B, and mouse embryonic stem (ES) cells treated with cell-permeant Cre (HTNCre) maintain a normal growth behavior employing Cre concentrations sufficient to induce recombination in more than 90% of the cells, whereas continuous application of high doses resulted in markedly reduced proliferation. HTNCre-treated ES cells maintain a normal karyotype and are still able to contribute to the germline. Moreover, we present an enhanced HTNCre purification protocol that allows the preparation of a concentrated glycerol stock solution, thereby enabling a considerable simplification of the Cre protein transduction procedure. The protocol described here allows rapid and highly efficient conditional mutagenesis of cultured cells.

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Metadaten
Document Type:Article
Language:English
Parent Title (English):Genesis
Volume:45
Issue:8
First Page:508
Last Page:517
ISSN:1526-954X
DOI:10.1002/dvg.20321
Publisher:Wiley-Blackwell
Date of first publication:2007/07/27
Departments, institutes and facilities:Institut für funktionale Gen-Analytik (IfGA)
Dewey Decimal Classification (DDC):600 Technik, Medizin, angewandte Wissenschaften / 610 Medizin und Gesundheit / 610 Medizin und Gesundheit
Entry in this database:2018/07/14

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