Study on the Colorimetric Detection for Mercury Ion Based on Nuclease and G-quadruplex DNAzyme

With the rapid development of industry, a great quantity of discard mercury has been discharged into and damaged the environment. By the statistics of United Nations Environment Programme(UNEP), China suffers the greatest mercury threat of the world. When organism ingests mercury by the way as drinking water, it can be accumulated rather than be expelled through the metabolism. With the accumulation of mercury, multisystem damage including nerve toxicity and renal toxicity imperils the health of biology. Owing to its high solubility and stability, Hg2+ion is the most usual found of mercury pollution and the high specificity and sensitivity detection of Hg2+in the water means significantly.As for the detection of mercury, methods including cold atomic fluorescence spectrometry, cold atomic absorption spectrometry, chromatography and inductively coupled plasma mass spectrometry have been widely used. Though these methods work well, they require sophisticated instrumentation and complicated sample preparation meanwhile, which restricts the spread in practical application. Recently, the discovery of specific thymine-Hg2+-thymine(T-Hg2+-T) structure attracts international concern. Plenty of Hg2+sensors have been designed based on the structure. Our research exactly employs T-Hg2+-T structure, along with the digestion ability of nuclease and excellent performance of nanoparticles to construct novel G-quadruplex DNAzyme based Hg2+colorimetric sensors. Follows are the main points of our paper.Chapter One:IntroductionIn this chapter, the basic concept of ion biosensor was introduced. And the application of T-Hg2+-T in Hg2+detection was reviewed in detail, together with the character and utilization of DNAzyme. Secondly, we presented the application of nanoparticles and nuclease hydrolysis in the biosensors to clarity the theoretical basis of our dissertation. At last, the purpose and significance of the research were pointed out.Chapter Two:A novel Hg2+-sensing assay based on G-quadruplex DNAzyme modified Au NP aggregations, MNPs and endonuleaseA specific optical strategy based on the amplification of DNAzyme bridged Au nanoparitcle(Au NP) aggregates, the separation of magnetic nanoparticles(MNPs) and endonulease digestion has been developed for trace mercury ions detection. When Hg2+was present, DNA modified magnetic capture nanoparticles(S1/MNPs) could hybridize with DNA coated Au capture nanoparticles(S2/S3/Au NPs) by forming thymine-Hg2+-thymine structure to compose the sensing assay. By adding crosslinking strand and DNAzyme modified Au NPs(S4/S5/Au NPs), aggregates were shaped to realize the amplification of signal. With the adding of EcoR V, dsDNA were digested and Au NP aggregates with DNAzyme were released to catalyze the colorless ABTS conversion into a blue-green product by H2O2-mediated oxidation. The detection limit for Hg2+was0.8nM, which was lower than the10nM US EPA limit in drinking water.Chapter Three:A novel sensing assay for Hg2+based on G-quadruplex DNAzyme and exonucleaseA sensitve optical strategy based on the amplification of two DNAzyme sequences on one DNA probe strand has been developed for mercury ions detection. One of the DNAzyme sequences was designed on the3’-terminal of probe, which was rich of thymine and had both the target recognition and signal amplification ability. When Hg24existed, the dual-function sequence could catch the target to form the specific thymine-Hg2+-thymine structure and protect the whole probe from the digestion of adding exonuclease. While hemin was added, this sequence preferred to destroy the thymine-Hg2+-thymine structure to form G-quadruplex, and then catalyze the colorless ABTS conversion into a blue-green product by H2O2-mediated oxidation. Owing to the design of two DNAzyme sequences, the detection limit was3nM.