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Ability of siRNAs to knockdown recombinant HeV-luciferase and cause immune stimulation.

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posted on 2013-05-14, 00:54 authored by Jana L. McCaskill, Glenn A. Marsh, Paul Monaghan, Lin-Fa Wang, Timothy Doran, Nigel A. J. McMillan

HeLa cells were transfected with 40 µg/ml Poly IC, 40 nM control or HeV targeting siRNAs. siLuciferase was a positive control (grey bar), while scrambled (ScrM7) siRNA and Oligofectamine were negative controls (black bars). A) Transfection media was replaced after 4 hours and cells were infected with recombinant HeV-luciferase at an MOI of 0.5. Infected cells were incubated for 24 hours before luciferase activity was measured. Levels of luciferase luminescence were normalized to Oligofectamine control levels and are the mean ± S.D. of six biological replicates from two independent experiments. B) Cells were incubated for 12 hours before RNA extraction. Gene expression levels of IL-6 were quantified by qRT-PCR as relative expression to B-actin housekeeping gene. Results are the mean ± S.D. of six biological replicates from two independent experiments. Significant differences between Oligofectamine control and siRNAs are indicated (**p = <0.01, ***p = <0.001, ****p = <0.0001; two-sided t-test).

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