Date of Award

2024

Document Type

Open Access Master's Thesis

Degree Name

Doctor of Philosophy in Chemistry (PhD)

Administrative Home Department

Department of Chemistry

Advisor 1

Shiyue Fang

Advisor 2

Yinan Yuan

Committee Member 1

Ashutosh Tiwari

Committee Member 2

Lanrong Bi

Committee Member 3

Yunhang Hu

Abstract

Synthetic long oligodeoxynucleotides (ODNs) have found wide applications in diverse fields such as chemical biology, synthetic biology, and genes and genomes synthesis. Those applications leading to a significant demand for their production. However, traditional methods for purifying synthetic ODNs present notable drawbacks, particularly their inability to purify long ODNs, rendering long ODN synthesis challenging. To address these issues, chemical synthesis techniques for long ODNs coupled with non-chromatographic purification methods have been developed. This technology makes the purification of long synthetic ODNs feasible and offers a viable pathway for producing genes and genomes with extensive repeats or stable secondary structures, which are either unattainable or exceedingly complex to achieve using conventional methods. Despite these advancements, the use of controlled pore glass (CPG) as a solid support for ODN synthesis poses its own limitations. In response, novel surface-functionalized solid support materials have been identified to mitigate these challenges. Direct synthesis up to 1,728-mer ODN on the surface has been reported. Results indicate that ODN synthesis on these surfaces eliminates steric hindrance and reduces errors commonly encountered with traditional CPG. Furthermore, while the catching-by-polymerization (CBP) method has proven effective, the affordability of polymerizable tagging phosphoramidites (PTPs) remains a bottleneck. To overcome this hurdle, scalable synthesis methods for the four nucleoside precursors have been discovered, offering chromatography-free alternatives and making the CBP method more efficient and cost-effective.

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