Synthesis Of Fluorescent DNA-templated Silver Nanoclusters And Label-free Detection Of DNA And Tumor Cells

Due to the excellent spectral and photophysical properties, fluorescent metal nanoclusters composed by several tens of atoms, has received widespread concerns. Currently, fluorescent metal nanoclusters based fluorescent assays have been widely used in biochemical analysis and biosensors, such as biochemical analysis and bio-sensors, owing to their convenience, sensitivity and ease of operation. Among them, the work using DNA as a template to synthesize fluorescent silver nanoclusters (AgNCs) aroused special attention and different silver nanoclusters with their fluo-rescence emission ranged from the visible region to near infrared region were pro-duced by changing the DNA sequence. Moreover, based on the inherent nucleic acids properties of template DNA, nucleic acid tool enzymes can also be introduce to this DNA-AgNCs-based sensing for signal amplification. The high binding capability of DNA has allowed the development of sensors for the detection of diferent molecules based on DNA-templated AgNCs. Consequently, on the one hand, we chose DNA-templated AgNCs as an effective signal indicator for label-free detection of DNA via combining nucleic acid tool enzymes-aided signal amplification; on the other hand, we hope to screen of a new DNA template for synthesis of AgNCs with high brightness and stability, and then apply them for biochemical analysis. The main research contents are as follows:1. Label-free detection of DNA based on DNA-templated fluorescent silver nanoclusters and exonuclease ?-assisted signal amplification strategiesIn recent years, DNA-templated AgNCs as a new type of fluorescence signal element has been widely studied and applied in analytical sensing field. The concen-trations of template DNA for synthesis of AgNCs is at micromole, which restricts its practical application for the detection of trace level targets. Herein, we presented a label-free DNA detection method based on DNA-templated AgNCs and exonuclease ?-assisted signal amplification. In this strategy, the AgNCs-forming template was designed to hairpin structure (denoted as HP), which can not as a template for sythe-sis of AgNCs directly due to its double-chain structure. Another 3'-protruding ter-minus of double-chain probe (denoted as HB) was designed to recognize target DNA. In the presence of target DNA, the P/HB probe could recognize it and formation of intact duplex DNA with blunt 3'-terminus. In this case, exonuclease ? could pref-erentially bond to the duplex region and catalyze the stepwise removal of mononu- cleotides from the blunt 3'-terminus. After the duplex was fully consumed, the target DNA and P were released. Then the released target DNA could bind to another 3'-protruding terminus of P/HB probe to trigger a new cleavage process. The released P could hybridize with HP hairpin to form DNA duplex with a blunt 3'-terminus, and then exonuclease ? could catalyze the cleavage the duplex DNA, releasing again the P for next cleavage process. Simultaneously, numerous single stranded AgNCs-forming sequences (T-DNA) were liberated and AgNCs could also be syn-thesized with high fluorescence signal. Thus, the concentration of target DNA could be identified by AgNCs1 fluorescence changes. This method provided a simple, iso-thermal, and low-cost approach for sensitive detection of DNA and holded a great potential for early diagnosis in gene-related diseases.2. DNA-templated formation of fluorescent silver nanoclusters with high stabil-ity and its application for tumor cells detetionDNA templated fluorescent silver nanoclusters were sequence-dependent, and various DNA sequences were selected to synthesize AgNCs with tunable fluores-cence emissions throughout the visible and near infrared region. As a development of DNA-templated AgNCs, we here reported a new template DNA, which could induce fomation of fluorescent AgNCs with high brightness and stability. In order to extend the application of this DNA-AgNCs, this template DNA was linked with an aptamer of tumor cell (sgc8c). This structure was used to form fluorescent AgNCs in the later stage and then to recognize the target tumor cells. The results showed that aptamer will not influence the luminescent property of DNA-AgNCs, and AgNCs-forming template will not affect the recognizetion capability of aptamer. It was demonstrated that the label-free aptamer strategy could efficiently and specially detect tumor cells. The result implied that this flurescent probe holds considerable potential for other targets detection.