G-Quadruplex Probes For The Label-Free Fluorescent Detection Of Heavy Metals And Telomerase

Heavy metal pollution is caused by heavy metals or their compounds in theenvironment. With the rapid development of industrialization process, heavy metalsare causing increasingly serious damage to the environment. As a kind of dangerouscontaminants, they are often accumulated in organisms but can’t get degraded. Heavymetal ions will do varying degrees of harm to the ecosystem, even at extremely lowconcentration. Pb2+, Hg2+are among highly toxic heavy-metal ions that are threateninghuman health. At present, chemical sensor technologies play a very important role inheavy metal ion detection. In the field of environmental monitoring and life science, itis of very important significance to detect these ions qualitatively or quantitatively.Telomerase is a ribonucleoprotein that binds to the telomere ends and stimulatesthe elongation of the telomeres. The telomerase-induced elongation of the telomerescompensates for the natural telomere shortening process, thus transforming the cellsinto immortal and malignant cells. Indeed, in over85%of cancer cells elevatedamounts of telomerase were found, and it is considered as a versatile biomarker forcancer cells. Thus, the sensitive detection of telomerase activity and its inhibition isimportant both for diagnostics and for the screening of anticancer drugs.According to the reported literatures, this thesis developed several novellabel-free fluorescent biosensing methods for the detection of Pb2+, Hg2+andtelomerase. The detailed methods are described as follows:In Chapter2, using K+and Pb2+as two inputs, we develop a novel label-freeINHIBIT logic gate for Pb2+detection. A label-free molecular logic system using aconformational switch from K+mediated G-quadruplex complex with NMM to Pb2+mediated G-quadruplex complex is developed. Under this design, we find that thePb2+-stabilized G-quadruplex is more stable compared to a K+system and Pb2+can infact displace K+to form a Pb2+-stabilized G-quadruplex. Based on the fact that theK+-stabilized G-quadruplex is sensitive to Pb2+, we design the logic system as a Pb2+sensor, this method is simple in design, fast in operation and possesses high sensitivityand selectivity.In Chapter3, we found that for G-rich DNA sequences with many T loopresidues the Hg2+-stabilized T-Hg-T is more stable than ThT induced G-quadruplexsystem. Therefore, Hg2+can specifically induce the formation of duplex nucleic acidDNA structures from DNA G-quartets. Based on the this fact, a highly sensitive label-free sensor for Hg2+was designed by inhibiting ThT as DNA G-quadruplexesfluorescent inducer. We believe that the present reported Hg2+sensor will provide aversatile tool for the determination of Hg2+in environmental samples. This assay alsoprovides a highly sensitive and specific sensing platform for Hg2+bioactivity analysis.In Chapter4, we presented a novel biosensing technology for the detection oftelomerase activity in cancer cells based on the fact that guanine can triggertransformation of AgNCs from a dark species to a bright red-emitting species. In thisassay, the primer contains oligonucleotides as scaffolds for the synthesis of AgNCs.This primer can act as scaffolds for the synthesis of dark AgNCs without telomerase.The bright red-emitting clusters AgNCs can be obtained when the primer addshexameric repeats (TTAGGG)n by the action of telomerase.The results indicated thatthe method could be used for sensitive determination of telomerase in a concentrationrange from500to50000HeLa cells. Given the simplicity, convenience of thisapproach, the proposed method may provide an alternative approach for the study ofthe telomerase activity.