RNA interference mediating small RNA molecules

A method for reducing the level of a target mRNA in a cell in vitro, comprising contacting a cell with an isolated synthetically prepared, double-stranded RNA molecule which mediates a reduction in the level of a target mRNA, wherein said molecule consists of two separate RNA strands each independently consisting of 19 -25 nucleotides, wherein said separate RNA strands form a single continuous double-stranded region and wherein at least one strand of said RNAi molecule has a 3 ′-overhang from 1 -5 nucleotides, wherein one of said strands is complementary to a portion of a target mRNA, and said RNAi molecule comprises at least one stabilizing modification at the 5 ′ end or the 3 ′ end, or both ends, of the double-stranded RNAi molecule, wherein said stabilizing modification is selected from the group consisting of a sugar modified ribonucleotide and a backbone-modified ribonucleotide containing a phosphorothioate group, wherein said sugar modified ribonucleotide 2 ′-OH group is replaced by a member selected from the group consisting of H, OR, R, halo, SH, SR, NH2, NHR, NR2 and CN, wherein R is C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, and halo is F, Cl, Br or I.

A method for reducing the level of a target mRNA in a cell in vitro, comprising:

• contacting a cell with an isolated synthetically prepared, double-stranded RNA molecule which mediates a reduction in the level of a target mRNA, wherein said RNA molecule consists of two separate RNA strands each independently consisting of 19-25 nucleotides, and
• wherein at least one strand of said RNA molecule comprises a 3 ′-overhang from 1-5 nucleotides, wherein one of said strands is complementary to a portion of a target mRNA, and said RNA molecule comprises at least one stabilizing modification at the 5 ′ end or the 3′ end, or both ends, of the double-stranded RNA molecule, wherein said stabilizing modification is selected from the group consisting of a sugar modified ribonucleotide and a backbone-modified ribonucleotide containing a phosphorothioate group, wherein said sugar modified ribonucleotide 2′-OH group is replaced by a member selected from the group consisting of H, OR, R, halo, SH, SR, NH2, NHR, NR2 and CN, wherein R is C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, and halo is F, Cl, Br or I.

A method for reducing the level of a target mRNA in a cell in vitro, comprising:

• contacting a cell with an isolated synthetically prepared, double-stranded RNA molecule which mediates a reduction in the level of a target mRNA, wherein said RNA molecule consists of two separate RNA strands each independently consisting of 19-23 nucleotides, and
• wherein at least one strand of said RNA molecule comprises a 3′-overhang from 1-3 nucleotides, wherein one of said strands is complementary to a portion of a target mRNA, and said RNA molecule comprises at least one stabilizing modification at the 5′ end or the 3 ′ end, or both ends, of the double-stranded RNA molecule, wherein said stabilizing modification is selected from the group consisting of a sugar modified ribonucleotide and a backbone-modified ribonucleotide containing a phosphorothioate group, wherein said sugar modified ribonucleotide 2′-OH group is replaced by a member selected from the group consisting of H, OR, R, halo, SH, SR, NH2, NHR, NR2 and CN, wherein R is C1-C6 alkyl, C2-C6 alkenyl, C2-C6 alkynyl, and halo is F, Cl, Br or I.