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. 2009 May 27;17(11):1454–1462. doi: 10.1038/ejhg.2009.80

Figure 4.

Figure 4

Differential expression is abrogated with transfection, time in culture, and age of DNA. Represents the fold difference in transcriptional activity after normalizing luciferase RLUs/ to −308G/GL3Luc, which was set at 1. (a) Jurkat cells were transiently transfected with either −308G/GL3Luc or −308A/GL3Luc DNA and harvested after 24 h to assay for luciferase activity using the Dual-Luciferase reporter assay system (Promega). Transfections were performed using Lipofectamine 2000 (Lipid-mediated; Invitrogen) or by electroporation as indicated. The experiment was performed thrice in triplicate. (b) Jurkat cells were cultured for 1, 2, or 3 weeks before transfection. The experiment was performed in triplicate. (c) (i) Represents three consecutive experiments performed in triplicate using DNA prepared from the QIAfilter Endofree Plasmid Maxi Kit (Qiagen) earlier and stored at −20°C. (ii) Represents the next three consecutive experiments performed in triplicate using fresh unstored DNAs prepared from the QIAfilter Endofree Plasmid Maxi Kit. The average fold differences of −308A/GL3Luc over −308G/GL3Luc from all experiments after normalization of raw luciferase RLU/s values to −308G/GL3Luc are shown. Statistical significance was determined by Student's unpaired t-test. P<0.05 was considered significant.