| Heavy metals have been popular used in modern industry, agriculture, medicine and other fields. The problem of heavy metal pollution is becoming more and more serious with the rapid development of science and technology. Heavy metals are difficult to degrade, they can be only one form into another form, eventually gathered in human body, endangering human body health. Therefore, detecting the heavy metal residues of agricultural products and the environment is particularly important. In this paper, some new methods for detection of heavy metals mercury, lead ion are designed in order to meet the demand for detection of heavy metals in daily life. Focus mainly involves the following aspects:1. The fluorescent sensor based on T- Hg2+-T structure for specificity detection of mercury ionIn chapter 2, a highly sensitive and selective method for detection of mercury ions was developed based on real-time fluorescent quantitative polymerase chain reaction(RT-q PCR) technology and the specific binding of Hg2+ in thymine-thymine mismatch complexes(T-Hg2+-T). The signal for detection of Hg2+ was generated using the RT-q PCR amplification of a template strand of DNA. The sensor exhibited an excellent linear response between cycle threshold(Ct) and mercury ion concentration in the range of 0.05–100 n M. Under optimized conditions, the detection limit of this method for aqueous Hg2+ was as low as 30 p M. In addition, this system showed good selectivity for Hg2+ over other metal ions. This new approach can be used in the analysis of Hg2+ in water and other samples.2. The new method Based on the GR- 5 DNA enzymes for detection of lead ionIn chapter 3, We report a facile approach for realize accurate detection of lead ion based on real-time fluorescent quantitative polymerase chain reaction technology and a lead-dependent DNAzyme, termed GR-5. In this method, substrate DNA is cleaved at the site of adenosine ribonucleotide by the GR-5 DNAzyme in the presence of lead ion, resulting in a decrease in template DNA available for PCR and a consequent change in signal detection(cycle threshold(Ct) value). This novel approach takes advantage of the exponential amplification of PCR and the specific recognition of the GR-5 lead-dependent DNAzyme to provide Pb2+-specific detection with an excellent linear relationship between Ct value and Pb2+ concentration from 1–500 n M. The correlation coefficient of the standard curve was 0.99 and the limit of detection was 0.7 n M. Moreover, the above sensory system showed good selectivity for Pb2+ over other metal ions.3. The new method based on exonuclease Ⅲ fluorimetry ultrasensitive detection of mercury ionIn chapter 4, a new simple and rapid biological sensor for detection of mercury ions in aqueous solution is developed. The sensor is based on the exponential of real-time fluorescent quantitative PCR amplification and quantitative, combined with exonuclease Ⅲ can shearing the 3’ end of closed double-stranded DNA. In presence of mercury ions, template DNA combining with complementary DNA by the specific binding of Hg2+ in thymine-thymine mismatch complexes(T-Hg2+-T) is formed the 3’ end of closed double-stranded DNA and then template DNA is cut by exonuclease Ⅲ. The amount of template DNA for amplification is affected by the amount of added mercury ions. The concentration of mercury ions finally was reflected by the Ct values of real-time fluorescent quantitative PCR experimental result and realize to qualitative analysis. The linear range for detection of Hg2+ was 0.05–20 n M and the limit of detection(LOD) was as low as 0.03 n M. The recoveries for the tap water samples were in the range of 94%–105%, and the RSDs were less than 4.65%. This sensor can be used in the detection of mercury ion concentration in water samples. |
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