Date of Award
Doctor of Philosophy (PhD)
Peng George Wang
Post-synthesis DNA modification is a very useful method for DNA functionalization. This is achieved by using a modified NTP, which has a handle for further modifications, replacing the corresponding natural NTP in polymerase-catalyzed DNA synthesis. Subsequently, the handle can be used for further functionalization, preferably through a very fast reaction. Herein we describe polymerase-mediated incorporation of trans-cyclooctene modified thymidine triphosphate (TCO-TTP). Subsequently, the trans-cyclooctene group was reacted with a tetrazine tethered to other functional groups through a very fast click reaction. The utility of this DNA functionalization method was demonstrated with the incorporation of a boronic acid group and a fluorophore. The same approach was also successfully used in modifying a known aptamer for fluorescent labeling applications. Boronic acid modified DNA molecules were further applied in aptamer selection for cancer cell recognition.
The second project was focused on developing fluorescent probes/sensors for biothiols. Because of the biological relevance of thiols and sulfides such as cysteine, homocysteine and hydrogen sulfide, their detection has attracted a great deal of research interest. Fluorescent probes are emerging as a new strategy for thiol and hydrogen sulfide analysis due to their high sensitivity, low cost, and ability to detect and image thiols in biological samples. A sulfonyl azide-based fluorescent probe has been developed for the quantitative detection of H2S in aqueous media such as phosphate buffer and bovine serum. In addition, another novel fluorescent probe has been developed for the detection of homocysteine. The fluorescence response is selective for homocysteine over other biologically abundant thiols such as cysteine and glutathione. In addition, a linear calibration curve was also be obtained for quantitative analysis in phosphate buffer and plasma. These selective fluorescent probes could be very useful tool for biothiol tests.
Wang, Ke, "Studies on Boronic Acid-modified Nucleotides for Diagnostic Applications and Development of Fluorescent Chemoprobes for Molecules of Biological Importance." Dissertation, Georgia State University, 2015.