Search

The use of nucleic acid technology has been of great importance in life science research. In several clinical trials patients with genetic disease are treated with help of innovations based on oligonucleotide therapy. The use of it is unfortunately limited in efficacy due to e.g., variable stability of oligonucleotides in biological fluids and in particular poor delivery to the site of action. Numerous modifications, introduced to functionalize oligonucleotides improve their properties and help to overcome the limitation connected to their uptake. With this thesis, I present our development of novel bioconjugates and technologies for functionalization of oligonucleotides. In chapter 2 (paper I), the development of four different synthetic m3G-Cap (nuclear localization signals) constructs is described. We provide a protocol that allows the conjugation of these for derivatives to oligonucleotide using “click” reaction. As a result, novel bioconjugates equipped with constructs containing the m3G-Cap were synthesized for the investigation of nuclear delivery. Chapter 3 (paper II) describes the synthesis of two biotinylated linkers that can be conjugated to both peptides and oligonucleotides for labeling purposes. The multi-step synthesis of the linker is followed by conjugation to oligonucleotides that was carried out on solid support as well as to a peptide in solution. In chapter 4 (paper III), the research presented in paper I and II is combined to allow the visualization of m3G-Cap action. The biotin linkers were also conjugated with similar Cap constructs containing modifications of the triphosphate bridge. All bioconjugates were evaluated in cell assays for uptake and splice-switching and the results show that the minimal m3G-Cap-biotin construct should contain a trinucleotide in order to act as a nuclear transport signal. Chapter 5 gives an insight into unpublished data on the development of a linker that enable multiple functionalization of oligonucleotides (ONs