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This helped the company extract the universal keys used to scramble target files.
To rule out the possibility of less favorable transcription caused by introduction of a long repeat sequence into the 3′UTR, we constructed a control plasmid optHRP-luc-ScrT by replacing the miR-199a-5p miR-199a-5p miR-199a-5p scramble target sequencesof the same length, withh wascrambleed based on targetck of sequenceant similarity tofany known miRNA (thele 1).
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As a comparison, the insertion of scramble targeting sequence did not repress transgene expression under normoxic condition significantly.
A control scramble targeting sequence (ScrT) of the same size was designed based on the lack of significant similarity to any known miRNA and subcloned into the same region (Table 1).
Inhibition was sequence-specific, since a scrambled control oligonucleotide or mismatched or scrambled target sequences failed to inhibit CAT expression.
Luc-CTL contains three random scrambled target sites that do not match any known miRNA sequence.
A negative control siRNA to a scrambled target in the DRAK2 gene (without sequence homology to any other known transcript as verified by BLAST analysis) was used in the mock transfection controls.
Another 20 scrambled prime facial target pairs (10 scrambled prime fearful targets, 10 scrambled prime neutral targets) were included to be new fearful and neutral faces.
Using ImageJ software, differential gel band intensities of scramble and target siRNAs were detected.
Control scrambled and target NRP-1 siRNA knockdowns resulted in comparable basal levels of unstimulated PDGFRβ phosphorylation.
As a control for off-target effects, non-specific scrambled siRNA (target sequence: 5′-ggacuaauaguugugcuccaauuua-3′) (Invitrogen) was used.
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