Development of oligonucleotides-based inhibitors and studies of the topological phenomenon of mobius DNA

• Main Author: Ng, Magdeline Tao Tao
• Other Authors: Li Tianhu
• Format: Theses and Dissertations
• Language: English
• Published: 2015
• Subjects: DRNTU::Science
• Online Access: http://hdl.handle.net/10356/62168
• Institution: Nanyang Technological University

With the aim of exploring new properties and applications of DNA enzymes such as Topoisomerase I and DNA ligase, during this study, the discovery of (1) oligonucletides-based DNA ligase inhibitors and (2) Topological phenomenon of Topoisomerase I and Mobius circular DNA were achieved. In Chapter 2, design and synthesis of a series of newly designed oligonucleotides containing a phosphorothioate group at the nick location. Our studies demonstrated that this modified phosphorothioate bond at the nick site indeed plays a crucial role in the inhibitory action on the activity of T4 and E.Coli DNA Ligase as it displays an IC50 value of 50nM and 46nM respectively, which is much more efficient than those reported in literature by Giovanni Ciarrocchi17 that has IC50 in micromolar range. In Chapter 3, we speculate besides the traditional method (using ethidium bromide based Topoisomerase I relaxation method) reported by Walter Keller et al8 developed 30 years ago, the superhelical density of circular DNA could be detected by manipulating the reaction times of Topoisomerase I in the absence of ethidium bromide. Therefore, in this study we report our examination on a straightforward method that does not require the addition of ethidium bromide. Our proposed method similarly gives the same result. Since Topoisomerase I may alter the topology of the DNA, we constructed the Mobius circular DNA using the designed oligonucleotides and studied topological phenomenon of the Mobius circular DNA.