Journal of General Virology, Vol 80, 15-21, Copyright © 1999 by Society for General Microbiology

ARTICLES

Construction and characterization of murine neuroblastoma cell clones allowing inducible and high expression of the prion protein

O Windl, H Lorenz, C Behrens, A Romer and HA Kretzschmar
Institut fur Neuropathologie, Universitat Gottingen, Germany.

A tetracycline-inducible expression system has been established for the prion protein (PrP) in murine neuroblastoma cells (N2a). For this purpose, N2a cells were first stably transfected with either the tetracycline-controlled transactivator or the reverse transactivator. After selection of N2a clones which carried one of these transactivators, the murine PrP gene (Prnp) was introduced under the control of the transactivator-responsive promoter in a second round of stable transfection. Stably double-transfected N2a clones carrying the reverse type but not the normal transactivator were found to be fully inducible, giving a low background of Prnp expression before induction and high expression after induction. Stably double-transfected N2a cells were at least as productive as N2a cells over-expressing Prnp permanently under the control of a strong viral promoter. Furthermore, the selected N2a clones allowed the Prnp expression level to be quantitatively controlled by varying the level of the effector substance, the tetracycline-derivative doxycycline. The clones were fully controllable, as over-expression could be switched on and off as desired. These N2a clones may become an important tool for elucidation of the cellular function of PrP and may pave the way for the tetracycline-inducible expression of many genes in this neuroblastoma cell line.

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