MicroRNA mediated knockdown of both E1A mRNA and protein measured by RT-QPCR and western blot after intravenous injection of 2×10¹⁰ vp of Ad5mir122.

<p>A) RT QPCR for the 13S E1A mRNA transcript in the livers of mice 48 hrs after intravenous injection with 2×10¹⁰ vp of Ad5mir122, Ad5WT, Ad5Luc or PBS. Ad5mir122 shows significantly reduced E1A mRNA during liver infection when compared to Ad5WT (N = 3). Statistical analysis was performed using a two tailed student T-Test (*P = <0.05). B) Western blot to confirm that all E1A proteins variants are knocked down. Each lane represents protein extracted from an individual mouse. Control lanes containing liver from mice treated with either an E1A deleted Ad5Luc vector or PBS show no E1A signal. Ad5WT treatment shows 3 clearly defined bands corresponding to proteins produced from the 13S (36 kDa), 12S (26 kDa) and a smaller fainter band which may represent the 11S or the 10S E1A transcript product. Treatment with Admir122 shows significant knockdown in E1A protein levels for all splice variants. The blot was exposed for 1, 5 and 10 minutes with the 5 minute exposure presented here. Molecular weights were calculated against a dual colour molecular weight ladder (Bio-Rad).