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Publication type: Baccalaureate Thesis

Year: 2013

Author(s): St√ľcklschwaiger, David

Title: Expression of the Murine Cationic Amino Acid Transporter 1 (mCat1) in HEK 293T cells.

Other title: Expression des murinen kationischen Aminosäure Transporters 1 (mCat1) in HEK 293T Zellen

Source: Bakkalaureatsarbeit, Vet. Med. Univ. Wien, pp. 29.


Fink Dieter

Hlavaty Juraj

Vetmed Research Units:
Institute of Laboratory Animal Science

Graduation date: 26.07.13

Introducing exogenous genes into living cells or tissues is important for the study of biologi-cal systems. Virus mediated gene transfer using retroviruses will lead to a stable integration and expression of the gene of interest. Because of their ability to stably integrate into the host genome they have been used to produce transgenic animals (LOIS et al., 2002). Lentiviral vectors packaged with the murine ecotropic envelope can only transduce cells expressing the murine retrovirus receptor mCat1 (MASUDA et al., 1999), increasing the biosafety of the lentiviral vectors used. For this study I cloned an mCat1-P2A-mCherry construct that leads to stoichiometric expres-sion of the murine retrovirus receptor mCat1 and the fluorescent protein mCherry. The self-cleaving short peptide sequence P2A links the expression of mCat1 and mCherry and allows us to determine the relative amount of the receptor by analyzing the fluorescent protein (KIM et al., 2011). The construct was inserted into a pCAG backbone and tested in HEK 293T cells. Only mCat1 expressing cells were receptive to the lentivirus packaged with the murine ecotropic enve-lope, which supports the assumption that such viral vectors can be used with minimal risks for researchers and the environment.

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