Cloning and Overexpression of AXL

pDONR223-AXL was obtained from Addgene, Cambridge, MA, USA (Addgene plasmid # 23945) [33 (link)]. AXL cDNA was cloned into the pLenti-C-mGFP (OriGene, Rockville, MD, USA) and SCC-25 cells were transduced with this vector or empty vector as control using lentiviral transduction. Fluorescence-activated cell sorting (FACS) of GFP+ cells (FACSAriaIII, BD Biosciences, Heidelberg, Germany) was used to select positive cells. Overexpression was subsequently estimated via Western blot.

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von Mässenhausen A., Brägelmann J., Billig H., Thewes B., Queisser A., Vogel W., Kristiansen G., Schröck A., Bootz F., Brossart P., Kirfel J, & Perner S. (2016). Implication of the Receptor Tyrosine Kinase AXL in Head and Neck Cancer Progression. International Journal of Molecular Sciences, 18(1), 7.

Publication 2016

pDONR223-AXL pLenti-C-mGFP FACSAriaIII

Cdna Cells Plasmid Vector Western blot

Corresponding Organization : Research Center Borstel - Leibniz Lung Center

Other organizations : University Hospital Bonn

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Variable analysis

independent variables

Transduction of SCC-25 cells with pLenti-C-mGFP vector containing AXL cDNA or empty vector

dependent variables

Overexpression of AXL protein (estimated via Western blot)
Selection of GFP+ cells (by FACS)

control variables

SCC-25 cells transduced with empty vector

controls

Positive control: SCC-25 cells transduced with pLenti-C-mGFP vector containing AXL cDNA
Negative control: SCC-25 cells transduced with empty pLenti-C-mGFP vector

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