Locked nucleic acid (LNA) oligomers were found to be very effective in their ability to modulate alternative splicing in vivo in transgenic mice that ubiquitously express a modified EGFP pre-mRNA containing an aberrantly spliced β-globin intron (IVS2-654). Following intraperitoneal injections, the splice-switching oligonucleotide LNA SSO-654 targeted to the aberrant 5′ splice site in EGFP-654 pre-mRNA corrected aberrant splicing and increased production of repaired EGFP mRNA mainly in the liver, colon, and small intestine. Little or no effect was detected in heart, lung, or kidney, the organ where most of the oligonucleotide was distributed after four consecutive daily injections. In the liver, LNA SSO-654 had an EC₅₀ of 3 mg/kg, approximately 17-fold more potent than its 2′-O-methyl congener. Moreover, in the liver, colon, and small intestine oral doses of 50 mg/kg resulted in detectable levels of splice switching. The effects of four daily injections at 25 mg/kg persisted for up to 29 days but did not result in liver toxicity. The results indicate that the LNA backbone confers sequence- and organ-specific functional biodistribution of the oligonucleotides and that these potent compounds have the potential to be safe and long-acting modulators of diseases treatable by splicing manipulation.