基于磁纳米颗粒的二段对称分裂式G-四分体DNA酶生物传感器用于Hg2+的快速检测

doi:10.6023/A14030234

摘要/Abstract

提出了一种可用于Hg²⁺快速检测的基于磁纳米颗粒与二段对称分裂式G-四分体DNA酶的生物传感器. 分别用紫外-可见光谱法, 圆二色光谱法和荧光显微镜成像技术对实验设计的DNA酶传感器进行了表征. 传感器中磁纳米颗粒的应用不仅可以直接从水样中通过磁分离方法分离和富集被测物Hg²⁺, 并且还能将游离的未与Hg²⁺结合的DNA酶和hemin等除去, 有效地提高检测灵敏度和降低背景信号; 此外, 二段对称分裂式G-四分体DNA酶的运用还可增强传感器的灵活性和选择性. 传感器对Hg²⁺检测的线性范围为0.8～20 nmol/L, 检出限为0.3 nmol/L. 当水体中的共存离子大量存在时, 传感器对Hg²⁺的检测仍具有高度特异性. 对实际水样的检测回收率在95.3%～104.4%之间. 实验设计的DNA酶传感器操作简便, 费用低廉, 具有良好的再生能力. 可用于天然水体和饮用水样品中痕量Hg²⁺的检测.

关键词: 磁纳米颗粒, 二段对称分裂式G-四分体, DNA酶, 生物传感器, 比色法, Hg²⁺

A biosensor composed of magnetic nanoparticles and symmetrically split G-quadruplex DNAzymes was proposed for the rapid and sensitive detection of Hg²⁺. It was mainly based on the formation of a special thymine-Hg²⁺-thymine (T-Hg-T) structure by T-riched nucleic acid sequences and Hg²⁺, and then G-quadruplex DNAzymes catalyzed the oxidation of H₂O₂ and 2,2'-azinobis-(3-ethylbenzthiazoline-6-sulphonate) (ABTS), so that the trace amount of Hg²⁺ in water can be successfully detected by colorimetry. The assembly of the biosensor was characterized by using of UV-Visible spectroscopy, CD spectroscopy and fluorescence microscopy imaging techniques respectively. The magnetic nanoparticles coated with T-riched short oligonucleotide (DNA1) via streptavidin-biodin interaction were employed as the capture elements. The long oligonucleotides (DNA2) consisting of the T-riched sequence in the middle and two G-riched fragments at the both ends were used as the sensing elements. The procedure of the protocol was divided into three steps. First: the magnetic nanoparticles covered with DNA1 (MNPs/DNA1) mixed with DNA2 were added into a water sample to capture the target Hg²⁺ by forming the special T-Hg-T structure. Incubating 2 h at 37 ℃, two G-riched fragments at the ends of DNA2 were close to each other assembling the symmetrically G-quadruplex DNAzymes in the presence of hemin. Then, the supernatant was discarded by magnetic separation, and the residues were washed several times to resuspend in the mixture solution containing definite amount of H₂O₂ and ABTS. The catalytic signal of absorbance of ABTS⁺ was recorded by UV-Visible spectrophotometer. The utilization of MNPs can greatly reduce the background signal, and the employment of symmetrically split DNAzymes made the design of the experiment more flexible and selectable. The biosensor was sensitive, and the calibration curve was identified in the range from 0.8 nmol/L to 20 nmol/L with a detection limit of 0.3 nmol/L. And the biosensor can detect Hg²⁺ without interference in the presence of a large number of competing ions. This sensor was simple, inexpensive, and renewable. Satisfactory values between 95.3% and 104.4% were obtained for the recovery experiments, and it can meet the requirement of detection of trace Hg²⁺ in natural water especially in drinking water.

Key words: magnetic nanoparticles, symmetrically split G-quadruplex, DNAzymes, biosensors, colorimetry, mercury ions

引用此文

张佳佳, 代佩卿, 李超, 李南忘, 程圭芳, 何品刚, 方禹之. 基于磁纳米颗粒的二段对称分裂式G-四分体DNA酶生物传感器用于Hg²⁺的快速检测[J]. 化学学报, 2014, 72(9): 1029-1035.

Zhang Jiajia, Dai Peiqing, Li Chao, Li Nanwang, Cheng Guifang, He Pingang, Fang Yuzhi. A Symmetrically Split G-quadruplex DNAzymes Biosensor Based on Magnetic Nanoparticles for the Rapid Detection of Hg²⁺[J]. Acta Chimica Sinica, 2014, 72(9): 1029-1035.

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基于磁纳米颗粒的二段对称分裂式G-四分体DNA酶生物传感器用于Hg²⁺的快速检测

A Symmetrically Split G-quadruplex DNAzymes Biosensor Based on Magnetic Nanoparticles for the Rapid Detection of Hg²⁺

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