The use of trademark symbols does not necessarily indicate that the name is trademarked in the country where it is being read it indicates where the content was originally developed. Products and content are covered by one or more patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc (NEB). Cells were photographed 48 hours post-transfection. GFP Silencing in COS-7 Cells: COS-7 cells co-transfected in a 24 well plate with a plasmid expressing GFP in the absence (control) or the presence of 30 ng (4 nM) of GFP siRNA prepared using the ShortCut RNAi Kit. Digests were analyzed by 20% TBE polyacrylamide gel electrophoresis. (B) dsRNA fragments (1 kb and 175 bp) were digested with ShortCut RNase III. Marker lane contains a mixture of 21 bp siRNA Marker and 100 bp DNA Ladder (NEB #N3231). Digests were analyzed by 20% TBE polyacrylamide electrophoresis. ShortCut RNase III digestion of dsRNA: (A) Varying amounts of ShortCut RNase III were incubated with 2 µg of a 500 bp dsRNA for 20 minutes at 37☌ in a 50 µl reaction. 1.5 units (1 µl) of ShortCut RNase III is sufficient to convert 1 µg of dsRNA into siRNA suitable for RNA interference in mammalian cells. ShortCut ® RNase III, used with its manganese-containing reaction buffer, converts long double-stranded RNA into a heterogeneous mix of short (18–25 bp) interfering RNAs (siRNA) suitable for RNA interference in mammalian cells (1–3).
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