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. 2015 Oct 30;46(1):167–177. doi: 10.1002/eji.201444970

Figure 2.

Figure 2

Transfection of MDDCs with miR‐650 pre‐miR in MDDCs reduces mRNA levels of ISGs. Top three transcripts regulated in MDDCs transfected with miR‐650 pre‐miR (see Table 1) were chosen for further validation. (A) Gene expression levels of CXCL10, IFIT2, and GCH1 were measured using qRT‐PCR, normalized to GAPDH and expressed relative to scramble control. qRT‐PCR measurements were performed in triplicate and data are shown as mean ± SEM and are pooled from three independent donors. (B) Schematic representation of the miR‐650:IFIT2 duplex with the miRNA seed marked in bold. (C) Dual luciferase reporter assay testing the responsiveness of pmiR‐Glo vectors containing CXCL10, IFIT2, or GCH1 3′UTR sequences to miR‐650 pre‐miR. The empty vector served as negative control. Firefly activity was normalized to Renilla and is expressed relative to control pre‐miRNA. Measurements were performed in triplicates or quadruplicates and data are represented as mean ± SEM from three independent experiments. (D) Table highlighting ISGs that were significantly regulated by miR‐650 pre‐miR on a transcriptional level, including fold changes and in silico target site information (b.s.: binding sites, FC: fold change). (A and C) Statistical significance was tested using a paired Student's t‐test (*p < 0.05).