Synthetic oligonucleotides offer interesting prospects for the control of gene expression but clinical applications have been severely limited by their poor bioavailability. Cationic lipids have been widely used for the delivery of charged oligonucleotide (ON) analogues but most of the commercial formulations are toxic and poorly stable in the presence of serum proteins. We have developed a DOGS/DOPE liposome formulation named DLS (for delivery liposomal system), that allows for the efficient nuclear delivery of negatively charged antisense ON analogues as monitored by fluorescence microscopy and by their ability to correct deficient pre-mRNA splicing, even in serum-supplemented cell culture. Uncharged DNA mimics such as peptide nucleic acids (PNA), or phosphorodiamidate morpholino (PMO) ON are particularly interesting for their high metabolic stability and affinity for complementary RNA targets but they cannot be delivered with cationic lipids. Cell penetrating peptides (CPP), such as Tat or penetratin, have been used widely as conjugates for the delivery of various biomolecules and might be appropriate for neutral ON analogues. However, entrapment within endocytic vesicles severely limits the efficiency of PNA delivery by CPPs in the absence of endosomolytic drugs, such as chloroquine. The conjugation of new arginine-rich CPPs to PNA allows efficient nuclear delivery in the absence of chloroquine as monitored in a splicing correction assay. Both strategies have their advantages but DLS-mediated delivery remains more efficient than CPP delivery for the nuclear targeting of splice correcting ON analogues in vitro.