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Maetzig, T.* ; Kuehle, J.* ; Schwarzer, A.* ; Turan, S.* ; Rothe, M.* ; Chaturvedi, A.* ; Morgan, M.* ; Ha, T.C.* ; Heuser, M.* ; Hammerschmidt, W. ; Baum, C.* ; Schambach, A.*

All-in-One inducible lentiviral vector systems based on drug controlled FLP recombinase.

Biomaterials 35, 4345-4356 (2014)

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Open Access Green möglich sobald Postprint bei der ZB eingereicht worden ist.

Abstract
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Site specific recombinases are frequently used as gene switches in transgenic animals where recombination is induced by drug treatment or by tissue specific recombinase expression. Alternatively, lentiviral gene transfer can be utilized for the genetic modification of a wide variety of cell types, albeit systems for tight control of transcriptional activity are scarce. Here, we combined lentiviral gene transfer and the development of a tightly drug-controlled FLP recombinase for the construction of "All-in-One" inducible gene expression systems. Tight control of FLP activity was achieved through N-terminal fusion with a FKBP12-derived conditional destruction domain and a C-terminal estrogen receptor binding domain making recombination dependent on the presence of Shield-1 and 4-hydroxytamoxifen. Exploiting the capacity of FLP to mediate excision and inversion, "All-in-One" lentiviral gene switch vector systems were generated where drug-induced recombination resulted in abrogation of FLP expression and subsequent overexpression or knockdown of the prototypical tumor suppressor phosphatase and tensin homolog PTEN. "All-in-One" vectors proved their functionality in a variety of hematopoietic cell lines, and primary murine bone marrow cells. Our new vector system thus combines the ease of lentiviral gene transfer and the power of site specific recombinases for analysis of gene function.

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