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Technology

This designed p-shRNA before cleavage is generated from a dumbbell DNA template consisting of a 21 to 29 base pair region encoding a siRNA sequence, with one single stranded loop containing one or more guanosine residues, and the other loop containing no guanosine residues. The p-shRNA is then treated with RNase T1 which specifically cleaves single stranded RNA at the 3’ ends of guanosine residues, to produce siRNA repeats connected by single stranded linkers at one end, with the other end accessible for efficient Dicer cleavage. The inventors have shown that this Dicer-cleavable p-shRNA demonstrates much greater silencing activity than unmodified p-shRNA in GFP-expressing HeLa cells, as well as significant reporter gene knockdown in SKOV3 and UCI101 ovarian cancer cells. This p-shRNA’s greater molecular weight and flexibility compared to siRNA can potentially allow more stable complexation using less delivery materials making it ideal for optimizing gene delivery vehicles.