Haynes Lab:Notebook/RNA-seq of PcTF Transfected U2OS & SK-N-SH cell lines/2014/12/19
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NOTE: Today's procedure is a repeat of the experiment on 12/16/2014, using a higher-concentration plasmid DNA source.
In order to introduce a new gene into the mammalian cancer cells, we need to make the cells competent and present them with a prepared plasmid containing our gene of interest along with a selectable marker. Lipofectamine is a tranfection reagent that produces positively charged liposomes that envelop the plasmids to be introduced to the cells and encourage passage through the negatively charged cell membrane into the cells.
To get your plasmid to the appropriate concentration, add 2.0μg to each 1.5mL microcentrifuge tube and then add nuclease-free water to bring the total volume to 20μL.
Transfected cells are in the upper incubator, third shelf from the top.