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Miah SM , Campbell KS
Expression of cDNAs in human Natural Killer cell lines by retroviral transduction
Methods Mol Biol. 2010 ;612 :199-208
PMID: 20033642    PMCID: PMC2798139   
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Abstract
Human NK-like cell lines are difficult to transfect using standard mammalian expression vectors and conventional transfection protocols, but they are susceptible to retroviral transduction as a means to introduce cDNAs. Our laboratory has exploited this technique to study a number of receptors in human NK cell lines. The method utilizes a bicistronic retroviral vector that co-expresses either drug resistance or enhanced green fluorescent protein (EGFP) in parallel with the gene of interest. After a single infection with recombinant retrovirus, transduced NK cells can be sorted for expression of EGFP or the transduced cell surface marker. Alternatively, cells expressing the transduced cDNAs can be selected for by treatment with neomycin, puromycin, or hygromycin. Using this method, the sorted/selected cells uniformly express the gene of interest and the expression is stable for many weeks of culture.
Notes
Miah, S M Shahjahan Campbell, Kerry S CA06927/CA/NCI NIH HHS/United States R01-CA083859/CA/NCI NIH HHS/United States R01-CA100226/CA/NCI NIH HHS/United States T32-CA009035/CA/NCI NIH HHS/United States Research Support, N.I.H., Extramural Research Support, Non-U.S. Gov't United States Methods in molecular biology (Clifton, N.J.) Nihms152823 Methods Mol Biol. 2010;612:199-208.