核酸适体识别荧光法检测汞离子

引用本文: 雷兆静, 张存政, 胡秋辉, 刘媛, 张强, 刘贤金. 核酸适体识别荧光法检测汞离子[J]. 分析化学, 2012, 40(12): 1827-1831. doi: 10.3724/SP.J.1096.2012.20127 shu

Citation: LEI Zhao-Jing, ZHANG Cun-Zheng, HU Qiu-Hui, LIU Yuan, ZHANG Qiang, LIU Xian-Jin. Aptamer-Based Fluorescence Assay for Hg²⁺ Determination[J]. Chinese Journal of Analytical Chemistry, 2012, 40(12): 1827-1831. doi: 10.3724/SP.J.1096.2012.20127 shu

核酸适体识别荧光法检测汞离子

雷兆静 ^1,2, ,
张存政 ^2, ,
胡秋辉 ^1, ,
刘媛 ^2, ,
张强 ^2, ,
刘贤金 ^{2,
,}

通讯作者: 刘贤金

基金项目:
本文系江苏省农业科技自主创新项目(No.CX(10)236) (No.CX(10)236)

苏州市科技支撑项目(No.SN201129) (No.SN201129)

中国博士后基金(No.20090451177) (No.20090451177)

江苏省博士后科研资助计划(No.0802023B)资助项目 (No.0802023B)

摘要: 通过对已知的Ni²⁺适体的改造,发现了对Hg²⁺有较强结合活性的核酸适体N1,基于N1的二级结构进行改造,获得了具有较高活性的核酸适体N5,并建立了荧光检测方法。以荧光基团FAM标记核酸适体,在94℃变性5 min,室温(25℃)下复性30 min后,适配体与标记有荧光猝灭基团DABCYL的一段互补序列Q2结合(适体与Q2的浓度比为1:3),加入系列浓度的Hg²⁺与互补序列竞争结合核酸适体,以荧光信号的变化定量分析Hg²⁺浓度。结果表明,N1与Hg²⁺结合呈线性,线性范围为1.25-20 mg/L;检出限为0.62 mg/L;以N1结构为基础改造合成的序列和结构均不同的适体N4,N5,N6,N7对Hg²⁺的识别结合活性均不同,其中适体N5与Hg²⁺结合活性最为灵敏,特异性高,线性范围为0.156-2.50 mg/L;检出限为78 μg/L。

关键词:

核酸适体
/ 荧光
/ 汞离子
/ 检测

English

Aptamer-Based Fluorescence Assay for Hg²⁺ Determination

1.
¹ College of Food Science and Technology, Nanjing Agricultural University, Nanjing 210095, China;

Corresponding author: 刘贤金

Received Date: 15 March 2012
Fund Project:
本文系江苏省农业科技自主创新项目(No.CX(10)236)

苏州市科技支撑项目(No.SN201129)

中国博士后基金(No.20090451177)

江苏省博士后科研资助计划(No.0802023B)资助项目

Abstract: A RNA aptamer modified to DNA (N1) that could bind to the target of Ni²⁺ was found that could bind specifically to Hg²⁺ ion with typical secondary structure. Thereafter, several DNA aptamers with a certain conformation of sequence mutation and splice were designed in order to increase the property of Hg²⁺ binding, and Aptamer-based fluorescence assay for the Hg²⁺ ion determination was developed. The addition of the Hg²⁺ to a mixture containing the duplex of a fluorophore labeled aptamer and a quencher-modified antisense nucleic acid (Q2) would force the release of Q2 from labeled aptamer, which was spontaneously accompanied by the increase of fluorescence intensity and made Hg²⁺ in quantitative analysis. It was shown that stable labeled aptamer DNA and Q2 hydrogen bonds formed at concentration ration of 1:3(labeled aptamer and Q2, respectively), under the condition of 5 min denaturation in 94℃ and 30 min renaturation in room temperature (25℃) in the fluorescence assay preparation. For aptamer N1, the fluorescence assay results showed that linear response toward Hg²⁺ concentration with fluorescence quenching system ranged from 1.25 mg/L to 20 mg/L, with the limit of detection 0.625 mg/L. In order to improve the sensitivity of aptamer N1, 4 oligonucleotides (N4, N5, N6, N7) were designed and synthesized based on the structure of N1. The results showed that N5 had the best affinity and specificity to Hg²⁺ which displayed the linear range for the Hg²⁺ concentration detection was 0.156 mg/L to 2.5 mg/L with a detection limit of 78 μg/L.

Key words:

Aptamer
/ Fluorescence
/ Mercury(Ⅱ)
/ Detection

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1.
沈阳化工大学材料科学与工程学院沈阳 110142

核酸适体识别荧光法检测汞离子

通讯作者: 刘贤金

English

Aptamer-Based Fluorescence Assay for Hg²⁺ Determination

Corresponding author: 刘贤金

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通讯作者: 陈斌, bchen63@163.com

中国化学会秘书处

核酸适体识别荧光法检测汞离子

通讯作者: 刘贤金

English

Aptamer-Based Fluorescence Assay for Hg2+ Determination

Corresponding author: 刘贤金

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