Xenbase Image ID: 76277

Fig. 1. In vivo GFP reporter activity of Xdcr- Mo-target constructs. Sequence alignment in the top panel (A) shows the two forms of Xdicer1 obtained by the analysis, with mis- matches indicated in white boxes. The target sequences of Xdcr-Mo1 and Xdcr-Mo2, which are the two Mos chosen for functional analysis, are shown in light blue and orange colours, respectively. Constructs carrying the GFP re- porter activity under the control of normal (WT) or mutated (MUT) Xdcr-Mo target sequences were cloned in pCS2 vector under CMV tran- scriptional control, as shown in (A). In the MUT constructs, the small letters indicate inserted mutations. (B) Scheme of the strategy followed to assay Xdcr-Mo1 ability to specifically inhibit its target. Red Fluorescent Protein (RFP) con- struct was always co-injected into early embryo as an internal standard, together with WT or MUT constructs, without (Control Mo1 target injection) or with (MutMo1; Mo1 target injec- tion) Xdcr-Mo1. (C) The ratio between GFP and RFP positive cells (n= number of cells analysed in three independent experiments) of mature embryonic retinas at st. 42. Bars indicate stan- dard error of the mean. Notably, only WtMo1 target injection (WtMo1) significantly decreases the ratio (p< 0.001), indicating the specificity of Xdcr-Mo1 to inhibit its specific target. Compa- rable results were obtained when injecting WtMo2 or MutMo2 constructs, with or without Xdcr-Mo2 (not shown). (D-F) Examples of reti- nas after Control Mo1 target injection without any Mo (D-D), MutMo1 target injection with Xdcr-Mo1 (E-E), or Mo1 Target injection with Xdcr-Mo1 (F-F). (D,E,F) RFP detection; (DEF GFP detection; (D,E,F) merge. Image published in: Decembrini S et al. (2008) Copyright © 2008. Reproduced with permission of the Publisher, University of the Basque Country Press. Larger Image Printer Friendly View

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