Synthesis of non-natural base-pairs : towards site-specific functionalisation of nucleic acids

Rights statement
Awarding institution
  • University of Strathclyde
Date of award
  • 2021
Thesis identifier
  • T16089
Person Identifier (Local)
  • 201566539
Qualification Level
Qualification Name
Department, School or Faculty
  • Alternative Splicing is one of the most important post-transcriptional processes as it facilitates the formation of multiple protein isoforms from the same gene sequence. Despite alternative splicing being highly regulated, anomalous splicing behaviour can be the root cause of disease states such as cancer. To date, research efforts have looked to explore and understand aspects of the spliceosome machinery but elucidating the effect of changes in the gene sequence upon splicing is significantly lacking. To define this critical effect, improved site-specific labelling of RNA is required. This thesis focuses on the synthesis and development of an orthogonal non-natural base pair system for the purpose of improving current RNA labelling strategies. Chapter one introduces alternative splicing, describes the regulatory processes involved and current methodologies for RNA labelling and their limitations. This chapter also introduces non-natural base pairs as a solution to overcome these limitations as well as other applications for non-natural base pairs. Chapter two describes the development of a library of guanosine analogues using a modular step efficient synthetic approach with the future aim of site specifically labelling viral RNA. Therefore this library was tested for antiviral activity and a trio of purine base analogues were tested for pairing fidelity opposite Hirao’s complimentary base to his purine analogues, the pyrrole analogue Pa by transcription assay. The antiviral testing was promising as no antiviral activity was found, however, the transcription assay did not show improvement over the Pa-s reference standard. Chapter three examines the utility of a new ribose hydroxyl protection strategy with the aim of achieving selective phosphoramidite synthesis of RNA nucleosides to improve the overall yield of non-natural purine phosphoramidites for solid phase RNA synthesis. Synthesis of a non-natural purine phosphoramidite precursor was achieved utilising the chosen strategy. Chapter Four describes the development of a modular synthetic route for 2′-deoxyadenosine phosphoramidite analogues, their incorporation into DNA by solid-phase DNA synthesis and an investigation of their utility for use as DNA aptamers. Incorporation of non-natural phosphoramidites into a 12mer DNA strand was achieved, however, the thermal stability test did not show improved pairing strength opposite Pa compared to the Ds reference standard.
Advisor / supervisor
  • Burley, Glenn
Resource Type
Date Created
  • 2020
Embargo Note
  • This thesis is restricted to Strathclyde users only until 6th December 2026.