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Title: The synthesis and study of oligo(deoxy)ribo-nucleotides and their analogues
Author: Ebel, Susanne
Awarding Body: University of Edinburgh
Current Institution: University of Edinburgh
Date of Award: 1993
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Laboratory procedures for the synthesis, workup and purification of oligoribonucleotides using RNA phosphoramidites protected with a) Fpmp- and b) TBDMS- groups on the 2'-hydroxl functions have been developed and employed in the following projects: 1. Modified minimised DNA/RNA ribozymes were synthesised and purified, which contained a) a dT4-loop, b) a hexaethylene glycol- or, c) a tetraethylene glycol-loop closing the catalytic site. Their catalytic activity was confirmed. The cleavage activity of the hexaethylene glycol minizyme was superior to that of the tetraethylene glycol minizyme, but cleaved less efficiently than the dT4-minizyme. 2. Tetra-biotinylated antisense RNA was synthesised and purified using two methods. The method using 2 successive additions of a multiple hydroxyl phosphoramidite followed by 1 addition of a single-addition biotin phosphoramidite, generated biotinylated antisense oligoribonucleotides of superior specificity in biotin/streptavidin based pre-mRNA depletion experiments. 3. The stability of tandem GA mismatches was analysed using thermal denaturation techniques. In DNA tandem GA mismatches of the kind 5' pry GA pu 3' adopted an amino-type basepairing and were very stable, tandem AG mismatches were imino-paired and significantly less stable. The amino-type base pair has a distinctly shifted phosphorus resonance in the 32P-NMR spectrum. Tandem GA mismatches in RNA did pair in the amino-form when flanked by 5' pyr..pu 3' but were highly unstable, as were tandem AG mismatches. Tandem GA mismatches in DNA/RNA hybrid sequences were also unstable. Multiple GA mismatches of the kind d(5' pyr GA pu 3') did not destabilise duplexes compared to T.A. base pairs. d(GA)n sequences formed homo-duplexes with exclusively G.A basepairing.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available