Generating Tetracycline-Inducible Cell Lines

cMYC and PIK3CA^H1047R (cloned from Addgene plasmid #12524) alleles were first cloned into pENTR4 no ccDB (686-1) (Addgene plasmid #17424) and subsequently shuttled into pLenti CMV/TO Puro DEST (670-1) (Addgene plasmid #17293) and pLenti CMV Hygro DEST (w117-1) (Addgene plasmid #17454), respectively (Campeau et al., 2009 (link)). Tetracycline-inducible cells were generated using pLenti CMV TetR Blast (716-1) (Addgene plasmid #17492). Lentiviral particles were generated using HEK293Tv (kind gift from Sam Benchimol) and MCF10A cell lines were transduced at the same time and with an equivalent viral titre.

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Lourenco C., Kalkat M., Houlahan K.E., De Melo J., Longo J., Done S.J., Boutros P.C, & Penn L.Z. (2019). Modelling the MYC-driven normal-to-tumour switch in breast cancer. Disease Models & Mechanisms, 12(7), dmm038083.

Publication 2019

pLenti CMV/TO Puro DEST (670-1) pLenti CMV Hygro DEST (w117-1) pLenti CMV TetR Blast (716-1)

Alleles Cell lines Cells Plasmid Tetracycline

Corresponding Organization :

Other organizations : Princess Margaret Cancer Centre, University Health Network, University of Toronto, Ontario Institute for Cancer Research

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Variable analysis

independent variables

CMYC allele
PIK3CA^H1047R allele

dependent variables

Not explicitly mentioned

control variables

Tetracycline-inducible cells generated using pLenti CMV TetR Blast (716-1) (Addgene plasmid #17492)
HEK293Tv (kind gift from Sam Benchimol) and MCF10A cell lines used for lentiviral particle generation and transduction

positive controls

Not specified

negative controls

Not specified

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