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. 2012 Sep 4;20(12):2304–2314. doi: 10.1038/mt.2012.158

Figure 7.

Figure 7

Aptamer RNA accumulation in viral, total, cytoplasmic, and nuclear RNA preparations. (a) Viral RNA was harvested from virus produced by transfection in the presence of aptamer. cDNA was synthesized from 200 ng of RNA and subjected to quantitative real-time-PCR using primers specific for 70N or 80N aptamers or reference RNAs (unspliced GAPDH for total RNA; spliced GAPDH for cytoplasmic RNA; U6snRNA for nuclear RNA; or human immunodeficiency virus-1 (HIV-1) gag for viral RNA). “No RT” controls were included for each sample, and each sample was assayed in triplicate. Each quantity was determined using the relative quantity (2–ΔΔCT) method where samples were first normalized to the corresponding reference RNAs, then to the pcDNA3.1 control (set to 1), and averaged. Values are shown as the mean ± SD. One-way ANOVA and Student's t-test were performed to determine statistical significance as compared to the original expression context (*P < 0.05; ** P < 0.01; ***P < 0.001). Experiments were performed three times, and a representative experiment is shown. (b–d) Total RNA and cytoplasmic and nuclear RNA from fractionated cells were collected from 293FT cells transfected with aptamers (b) 70.05, (c) 70.15, or (d) 80.80. cDNA, qPCR, and data analysis were performed as in (a) using the corresponding reference RNAs.