Highly functional short hairpin RNA

A fractured hairpin for use in gene silencing, the hairpin comprising:

• a first polynucleotide strand;
• and
• a second polynucleotide strand capable of forming a hairpin structure with the first polynucleotide that is capable of being processed by Dicer, the hairpin structure having from about 42 to about 106 nucleotides, the second polynucleotide strand comprising:
• a first region having at least 80% complementarity with the first strand and being capable of forming a first duplex region with the first strand;
• a second region coupled to the first region;
• a third region coupled to the second region;
• and
• a fourth region coupled to the third region and having at least 80% complementarity with the second region, the fourth region being capable of forming a second duplex region with the second region such that the third region forms a loop adjacent to the second duplex region.

A short hairpin RNA for use in gene silencing, the short hairpin comprising:

• at least one polynucleotide forming the short hairpin RNA having from about 42 nucleotides to about 106 nucleotides and being configured for being processed by Dicer, the hairpin RNA comprising:
• a first region;
• a loop region coupled to the first region;
• and
• a second region coupled to the loop region and being capable of forming a first duplex region with the first region;
• wherein at least one of the first or second regions includes at least two tandem nucleotides each having a 2′ modification such that processing by Dicer results in a sense strand having a first 5′ sense nucleotide at a first terminal nucleotide position and a second 5′ sense nucleotide at a second nucleotide position adjacent to the terminal nucleotide position, the first and second 5′ sense nucleotides having the 2′-modification.